Tag: Cytokine

Study reveals how SARS-CoV-2 hijacks lung cells to drive COVID-19 severity

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In a recent study published in the Journal of Experimental Medicine, researchers identified the cellular tropism and transcriptome consequences of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) by infecting human lung tissue and using single-cell ribonucleic acid sequencing (scRNA-seq) to rebuild the transcriptional program in “infection pseudotime” for distinct lung cell types.

Lower respiratory infections, such as coronavirus disease 2019 (COVID-19), are a leading cause of death worldwide, producing pneumonia and acute respiratory distress syndrome. Understanding their early phases is difficult. Researchers used classical histopathological approaches and single-cell multi-omic profiling to infer early phases in human pathogenesis from lung lavage, biopsy, or autopsy materials. These approaches reveal a thorough picture of COVID-19 pneumonia at unparalleled cellular and molecular resolution, implying infection models including alveolar epithelium, capillaries, macrophages, and myeloid cells.

Study: Interstitial macrophages are a focus of viral takeover and inflammation in COVID-19 initiation in human lung. Image Credit: Dotted Yeti / Shutterstock

About the study

In the present study, researchers developed an experimental COVID-19 model to investigate early molecular processes and pathogenic mechanisms of SARS-CoV-2 infection at the cellular level in native tissues of the human lung.

The researchers established SARS-CoV-2’s cellular tropism and its unique and dynamic impacts on host cellular gene expression in specific types of lung cells. Prominent targets were lung-resident macrophages, of which one SARS-CoV-2 takes over transcriptomes, inducing a targeted host interferon (IFN) antiviral program, and several chemokines and pro-fibrotic and pro-inflammatory and cytokines signaling to various structural and immunological cells of the lung.

To determine the early stages of COVID-19 in human lungs, the researchers sliced lung tissue obtained from surgical specimens or organ donor individuals into thick sections and used them for tissue culture analysis. Subsequently, they exposed the tissues to the SARS-CoV-2 USA-WA1 2020 strain at 1.0 multiplicity of infection (MOI) for two hours before allowing the SARS-CoV-2 infection to continue for two to three days. They performed a plaque test on culture supernatants.

The researchers separated the slices and examined them by scRNA-seq to evaluate host and viral genetic expression during the SARS-CoV-2 infection. They also examined the viral RNA molecules’ junctional structure and processing by analyzing the scRNA-seq dataset with the SICILIAN framework. They used molecular atlas markers to distinguish lung cell types in healthy lung slices and measure viral RNA levels in infected cells.

The team performed multiplexed single-molecule fluorescence in situ hybridization (smFISH) to confirm lung cell tropism findings and show infected cells. They used single-cell gene expression patterns to identify cellular targets for inflammatory and pro-fibrotic signals elicited by the SARS-CoV-2 infection of a-IMs. They devised a technique for purifying macrophage populations from human lungs with a SARS-CoV-2 spike (S) protein-pseudotyped lentivirus (lenti-S-NLuc-tdT) to investigate lung macrophage entrance routes.

The researchers productively infected human lung slices cultivated ex vivo with SARS-CoV-2, with production rising between 24 and 72 hours of culture. They heat-inactivated, ultraviolet (UV)-treated, or administered 10.0 µM remdesivir, an RNA-dependent RNA polymerase inhibitor used as a COVID-19 therapeutic, to prevent viral stock infection.

Results

The analysis showed that SARS-CoV-2 preferentially infects active interstitial macrophages (IMs), which can amass hundreds of SARS-CoV-2 RNA molecules, comprising >60% of the cell transcriptome and producing dense viral RNA bodies. Infected alveolar macrophages (AMs) exhibit no severe reactions, with spike (S) protein-dependent viral entrance into AMs utilizing angiotensin-converting enzyme 2 (ACE2) and the cluster of differentiation 169 (CD169) and IM entry via CD209.

They found canonical sub-genomic junctions between the unusual sequence reads beyond their 39 terminal regions, indicating canonical-type SARS-CoV-2 messenger RNA (mRNA) production in the pulmonary cultures. They also found hundreds of new subgenomic junctions, showing a wide range of non-canonical and canonical sub-genomic SARS-CoV-2 RNAs produced during pulmonary infection.

Model of initiation, transition, and pathogenesis of COVID-19 and the viral lifecycle in AMs and IMs. (a–d) Model of COVID-19 initiation in the human lung and transition from viral pneumonia to lethal COVID-19 ARDS. (a) SARS-CoV-2 virion dissemination and arrival in the alveoli. Luminal AM encounter virions shed from the upper respiratory tract that enter the lung. AMs can express low to moderate numbers of viral RNA molecules and can propagate the infection but “contain” the viral RNA from taking over the total transcriptome and show only a very limited host cell inflammatory response to viral infection. (b) Replication and epithelial injury. SARS-CoV-2 virions enter AT2 cells through ACE2, its canonical receptor, and “replicate” to high viral RNA levels, producing infectious virions and initiating viral pneumonia. (c) a-IM takeover and inflammation signaling. SARS-CoV-2 virions spread to the interstitial space through either transepithelial release of virions by AT2 cells or injury of the epithelial barrier, and enter a-IMs. Infected a-IMs can express very high levels of viral RNA that dominate (“take over”) the host transcriptome and can propagate the infection. Viral takeover triggers induction of the chemokines and cytokines shown, forming a focus of inflammatory and fibrotic signaling. (d) Endothelial breach and immune infiltration. The a-IM inflammatory cytokine IL6 targets structural cells of the alveolus causing epithelial and endothelial breakdown, and the inflammatory cytokines recruit the indicated immune cells from the interstitium or bloodstream, which flood and infiltrate the alveolus causing COVID-19 ARDS. Local inflammatory molecules are amplified by circulating immune cells, and reciprocally can spread through the bloodstream to cause systemic symptoms of cytokine storm. (e) Comparison of the SARS-CoV-2 viral lifecycle in AMs and IMs. Although both can produce infectious virions, note differences in viral entry receptors (AMs can use ACE2 and CD169/SIGLEC1, whereas IMs use CD209); viral RNA transcription of dsRNA intermediates (greater in AMs); replication of full-length genomic RNA (greater in IMs); viral takeover, formation of RNA bodies, and induction of a robust host cell inflammatory response (only in IMs), and cell destruction/death (only in IMs).

Heat, UV-C inactivation, or remdesivir therapy prevented the development of canonical and non-canonical connections. The team observed SARS-CoV-2 takeover of an activated IM subtype in 176,382 cells with high-quality transcriptomes obtained from infected lung slices of four donor lungs and in 112,359 cells from mock-infected slices (cultured without viral addition) and 95,389 uncultured control cells (directly from freshly cut lung slices). A differential gene expression study of a-IMs over infection pseudotime revealed host gene expression alterations corresponding to SARS-CoV-2 RNA levels.

The study found that COVID-19 pneumonia infection and takeover cause an early antiviral cell response specific to activated interstitial macrophages, resulting in a powerful immunological and fibrotic signaling center. Inflammasome activation is uncommon and only detectable late in a-IM infection. Blocking antibodies against CD169 and CD209 prevented entrance into IMs and AMs. The study also highlighted IMs as the most vulnerable lung target, with initial emphasis on inflammation and fibrosis. Two unique molecular lineages of macrophage targets react differently to SARS-CoV-2, influencing etiology and treatments.

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April 15, 2024
New insights into the exacerbation of psoriasis through specific genetic defects

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In a recent study featured in Nature Communications, researchers created mice that carry a gain-of-function (GoF) mutation in the gene encoding the inhibitor of nuclear factor kappa-b kinase subunit beta (IKBKB), known as the IKK2-encoding IKBKB gene. This was done to explore how this mutation works.

Study: IKK2 controls the inflammatory potential of tissue-resident regulatory T cells in a murine gain of function model. Image Credit: Gorodenkoff/Shutterstock.com

Background

Loss-of-function mutations demonstrate the importance of forkhead box P3-positive (Foxp3+) regulatory T cells (Tregs) in immunological control. Tregs mediate dominant tolerance and protect against autoimmune disorders.

They undergo positive selection in the thymus, and interleukin-2 (IL-2) protects them from apoptosis. Treg formation needs effective signaling downstream of the T-cell receptor (TCR), particularly the CARD11-BCL10-MALT1 (CBM) complex.

Mice lacking particular genes have a Treg deficit that causes a selective loss of cluster of differentiation 4-positive (CD4+) Helios+ thymic T cells.

Tregs move between lymphoid organs according to adhesion molecule expression. The presence of an activated or effector phenotype (eTreg) in recirculating Tregs increases disease risk.

About the study

The present study examined mice with an Ikbkb GoF mutation homologous to a problematic human IKBKB GoF variation.

The researchers tracked a cohort of mice with various Ikbkb genotypes and recorded the age at which skin disease appeared. Animal house technicians were unaware of the mouse genotype and identified abnormal Ikbkbmut/+ and Ikbkbmut/mut animals. The researchers examined the transcriptomes of tails and ears from Ikbkbmut/mut and Ikbkb+/+ mice.

The team investigated the inflammatory infiltrate in skin lesions and the nature of Treg growth inside pathological lesions. They created mixed bone marrow chimeras with allotype-marked donor cells from WT and mutant mice.

They isolated naïve CD4+ T cells from mouse splenocyte suspensions and activated them with Th17-inducing conditions. The researchers then counted IL-17+ Tregs ex vivo and labeled them for cytokine production after gating on Foxp3.

The researchers extracted them from WT mice and cocultured them with pure WT conventional T cells labeled with CTV to explore Tregs’ traditional immunosuppressive activity. They followed up with an in vivo test of mutant Treg suppression.

They analyzed mice for signs of systemic immunological dysregulation and created reciprocal bone marrow (BM) chimeras to study Ikbkbmut’s cell-intrinsic effects on the Treg phenotype.

The team obtained serum from recipient mice to analyze a panel of cytokines. They isolated green fluorescent protein (GFP)-labeled Foxp3+ Tregs from Ikbkbmut donors and implanted them into Ikbkbmut x Rag1−/− or IkbkbWT x Rag1−/− animals to establish disease cause as pro-inflammatory Treg activity.

The researchers used mice aged six weeks to 12 months for analysis. They performed flow cytometry, flow cytometric cell sorting, ex vivo PMA/ionomycin stimulation for cytokine production, T-cell polarization, an in vitro Treg suppression experiment, cell trace violet (CTV) labeling, and single-cell and bulk ribonucleic acid (RNA) sequencing studies.

Results

Canonical NF-κB overactivity led to the growth of pathogenic, NF-κB-dependent, and modified non-lymphoid tissue skin Tregs. Mice with Ikbkb GoF mutation heterozygosity developed psoriasis, and Ikbkb-mut mice included IL-17-producing Tregs.

These animals maintained suppressive function, indicating that normal CD4+ T cells are not the source of IL-17 in Ikbkb mutant mice. Foxp3+ CD4+ T cells from Ikbkb mutant mice maintained suppressive function.

The study additionally examined the effects of doubling the IkbkbGoF/GoF gene dosage on psoriatic arthritis, characterized by spondylitis, dactylitis, and distinctive nail abnormalities.

IkbkbGoF mice showed selective CD25+ and Foxp3+ Treg expansion, with a fraction expressing IL-17. These transformed Tregs were present in inflamed tissues, spleen, and blood, and their transfer was sufficient to cause illness without ordinary T lymphocytes.

Single-cell phenotyping and transcriptional investigations of isolated regulatory T cells indicated the non-lymphocytic tissue proliferation of Treg expressing Th17-associated genes, Helios, tissue-related markers such as CD69 and CD103, and a significant nuclear factor kappa B (NF-κB) transcriptome.

Overactive IKK2 caused dermal Treg accumulation and psoriasis. Heterozygous (Ikbkbmut/+) and homozygous (Ikbkbmut/mut) mutant mice developed skin illnesses with histopathological similarities to psoriasis.

Humans heterozygous for IKBKBV203I have combined immune insufficiency, but their Treg count increased. Ikbkbmut has a similar phenotype, with gene-dose-dependent lymphopenia caused by a decrease in αβ and γδ T cells in homozygous mice.

The study also found an increase in Th17 CD4+ T cells, strongly associated with psoriasis. Ikbkbmut/mut mice spleen Tregs produced more IL-17 than wild-type mice.

Interferon-gamma (IFNγ) production by Tregs was similar between WT and mutant animals, indicating that Ikbkbmut imparts an expansion of the IL-17-producing Foxp3+ Treg population.

Foxp3 deficiency and Treg functional abnormalities were associated with early-onset and severe widespread lymphadenopathy unrelated to the Ikbkbmut mutation.

Conclusion

The study linked psoriasis and psoriatic arthritis to NF-κB malfunction, which causes non-specific leukocytes to acquire an effector-like function, resulting in disease. The primary finding is a route that leads Foxp3+CD4+ tissue-resident Tregs to turn pro-inflammatory and pathogenic.

In vivo, a modified Treg population emerges owing to enhanced activity of the canonical NF-κB pathway. This route controls Treg abundance, increases tissue-resident Tregs, and mediates end-organ pathologies.

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March 27, 2024
Inflammatory responses fuel cardiovascular complications

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In a recent study published in the journal Circulation, researchers investigate the inflammatory response to acute respiratory distress syndrome (ARDS) within the heart.

Study: Virus-Induced Acute Respiratory Distress Syndrome Causes Cardiomyopathy Through Eliciting Inflammatory Responses in the Heart. Image Credit: Kateryna Kon / Shutterstock

The link between respiratory viral infections and CVD

Seasonal viral infections can range in severity from mild flu-like symptoms to potentially lethal ARDS. For example, despite being primarily a respiratory tract infection, coronavirus disease of 2019 (COVID-19) can lead to ARDS and other severe cardiovascular disease outcomes with high mortality rates.

Circulating immune cells may respond to COVID-19 by upregulating cytokine release, which can lead to myocardial injury. Cardiac macrophages, immune cells responsible for the myocardial inflammatory response, are increasingly being investigated for their role in ARDS. Recent evidence indicates that macrophage expansion, which can be accompanied by changes in the population size and relative abundances of various cardiac macrophages, is a characteristic feature of ARDS.

The main two types of cardiac macrophages include C-C chemokine receptor type 2 negative (CCR2^–) and CCR2⁺ macrophages. Further research is needed to determine the viral-induced contributions of these macrophages to adverse cardiac outcomes.

These data would allow clinicians to make informed intervention decisions and elucidate whether these outcomes are COVID-19-induced or if observed inflammation is a systemic immune response to viral infection. Furthermore, this information could support the development of future therapies to prevent cardiovascular disease (CVD) following recovery from COVID-19.

About the study

In the present study, researchers investigate the role of viral- and non-viral-induced ARDS-associated immune signals in altering cardiac macrophage populations, thereby impacting CVD parameters, including systemic inflammation.

This study was conducted at Massachusetts General Hospital and involved 33 control samples obtained from patients who died between September and December 2019, prior to the onset of COVID-19, as well as 21 samples obtained between May and July 2020 from patients who died from COVID-19-associated complications. Samples consisted of autopsy tissue excised from the left ventricular or septal region.

Simultaneously, in vivo studies involved a daily intratracheal administration of an ARDS cocktail of immunostimulatory agents to mice, which included resiquimod, imiquimod, lipopolysaccharide (LPS), and angiotensin-converting enzyme 2 (ACE2) inhibitor MLN-4760. This model allowed the researchers to reproduce clinical ARDS features in mice without the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2).

Patient data included results obtained from electrocardiogram (ECG), echocardiography, lung computed tomography (CT) scan, blood gas analyses, body temperature evaluation, bronchoalveolar lavage fluid (BALF) characterization, blood pressure measurements, and flow cytometry. Both human and murine autopsy samples were processed using ribonucleic acid (RNA) isolation, real-time polymerase chain reaction (PCR) assay, and enzyme-linked immunosorbent assays (ELISAs) for protein and gene expression determinations.

Similar immune responses in non-viral- and SARS-CoV-2-associated ARDS

In the absence of viral infection, mice treated with the ARDS cocktail exhibited significant weight loss over the five-day cocktail treatment period. This was accompanied by hypothermia, a common feature of both ARDS and septic shock, as well as a mortality rate of over 40% by day five.

Mice with ARDS exhibited bilateral opacities and immune cell infiltrations within their lungs, as well as reduced blood oxygenation. Furthermore, increased D-dimer, neutrophil, and monocyte levels were observed, as well as reduced blood pressure and lower heart rates in ARDS mice. Other inflammatory pathways that were activated in ARDS mice included increased levels of interleukin 6 (IL-6), IL-1ß, tumor-necrosis factor α (TNF-α), and interferon y (IFN-y), all of which are also associated with SARS-CoV-2 infection.

In both non-infected ARDS and SARS-CoV-2-infected mice, an increased infiltration of interstitial macrophages and reduced levels of alveolar macrophages were observed. Although both mouse models exhibited increased levels of cardiac macrophages, this immune response was more pronounced in infected mice. Nevertheless, both models’ subsets of cardiac macrophages were altered to similar levels.

Upon comparison of control and COVID-19 patient myocardium samples, SARS-CoV-2 infection recruited a more significant number of CCR2+ CD68+ macrophages, thus indicating that a robust immune response is elicited after severe infection compared to other life-threatening diseases.

“Our findings indicate that systemic and myocardial inflammatory signals elicited by virally induced ARDS may contribute to the cardiovascular complications and high mortality rates of this condition. In addition, our study confirms previous reports that SARS-CoV-2 infection increases overall macrophage numbers in hearts.”

The cardiac benefits of TNF-α immune therapy

TNF-α neutralizing antibodies were also administered to mice to evaluate their effects on immune activation during ARDS. To this end, TNF-α immune therapy reduced weight loss, improved body temperature, increased blood oxygenation, and led to better survival rates. Histological analysis indicated that ARDS mice receiving anti-TNF-α therapy exhibited reduced macrophages, Cxcl2, IL-1ß, and IL-6 expression within the lungs.

TNF-α therapy also improved systolic dysfunction, cardiomyocyte apoptosis, and monocyte infiltration in ARDS mice. Total cardiac macrophage counts and reduced expression of IL-1ß, IL-6, and TNF-α within the myocardium were also observed, thus demonstrating the anti-inflammatory benefits associated with TNF-α immune therapy in the lungs and hearts of mice with ARDS.

Conclusions

The study findings demonstrate that SARS-CoV-2 infection leads to significant alterations in cardiac macrophage subset levels, particularly increased levels of CCR2+ macrophages, in both mice and humans. Even in the absence of SARS-CoV-2 or another virus, the immune response to ARDS-like injury is capable of inducing significant alterations in heart macrophage levels, which may increase the risk of cardiovascular complications and mortality associated with ARDS.

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March 24, 2024
Certain genes slash severity and death risk in older men
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In a recent study published in The Journal of Infectious Diseases, researchers investigated the inflammation outcomes of three different Interleukin-1 receptor antagonist gene (IL1RN) single-nucleotide variants (SNVs) in acute severe respiratory syndrome coronavirus 2 (SARS-CoV-2) infection patients. Their retrospective study included almost 2,600 confirmed severe coronavirus disease 2019 (COVID-19) patients and showed that the IL1RN CTA haplotype and its rs419598 C/C SNV dramatically attenuated COVID-19-associated hyperinflammation, a characteristic of severe SARS-CoV-2 infections.

Observed outcomes were substantially improved in men compared to women, with men depicting 15% reduced mortality over women with the same SNV. These findings were most extreme for older men, with patients with the rs419598 C/C SNV above the age of 74 presenting 80% less mortality risk than their non-SNV-expressing age-matched counterparts. This study is one of the first to elucidate the genetic determinants of COVID-19 pathology and may form the basis for personalized future interventions against the disease.

Study: Interleukin-1 Receptor Antagonist Gene (IL1RN) Variants Modulate the Cytokine Release Syndrome and Mortality of COVID-19. Image Credit: Adao / Shutterstock

COVID-19 and the dangers of CRS

The coronavirus disease 2019 (COVID-19) represents one of the worst pandemics in human history, responsible for almost 7 million deaths worldwide and leaving hundreds of millions of survivors with long-lasting clinical symptoms. In severe cases, the acute respiratory syndrome coronavirus 2 (SARS-CoV-2) may result in multiorgan failure, acute respiratory distress syndrome (ARDS), and even death in 10-20% of affected patients.

Research has shown that severe COVID-19 symptoms are often associated with elevated plasma cytokine levels, especially those of interleukin 1β (IL-1β), IL-2, and IL-6. Unfortunately, a number of immunotherapy drugs, including those used to treat COVID-19, have been implicated in the overexpression of these ILs, a condition similar to cytokine release syndrome (CRS). Previous work by the present research group identified that IL1RN haplotypes containing the rs419598, rs315952, and rs9005 single-nucleotide variants (SNVs) could alter osteoarthritis and rheumatoid arthritis severity by attenuating hyperinflammation.

Unfortunately, the role of genetics in COVID-19 pathology remains poorly understood. The present study aims to shine a light on this knowledge gap by investigating the role of IL1RN SNP in moderate-to-severe COVID-19 infections.

About the study

Previous research by the current group identified the associations of IL1RN genetic variants with osteoarthritis and rheumatoid arthritis outcomes. It revealed that three SNVs (rs419598, rs315952, and rs9005) improved disease outcomes via hyperinflammation reduction mechanisms. The present study aims to investigate if the same genetic variants could improve COVID-19 outcomes due to the central role of hyperinflammation in severe COVID-19 pathology.

The study is a retrospective, observational study comprising data from adult (19+) patients admitted to Tisch Hospital, New York, United States, between March 2010 and March 2021. The cytokine profiles of these patients were compared against healthy age, sex, and body mass index (BMI)-matched controls without a clinical history of COVID-19 exposure. Real-time reverse transcriptase-polymerase chain reaction (RT-PCR) assays were used to confirm COVID-19 status and severity. Data sources comprised sociodemographic (sex, age, race, and ethnicity) and medical data obtained from hospital records and discarded COVID-19 blood samples (for plasma extraction). Data generation included whole-genome sequences (low coverage) of participants’ blood. The gencove.org database was used to annotate common SNV genotypes for each sequenced sample.

Three IL1RN genotypes, namely rs419598, rs315952, and rs9005, formed the focus of this study and were extracted from patients’ plasma samples during routine COVID-19 care. However, since multiple cytokines of interest were not included in routine care, plasma samples from 359 randomly selected study participants and their demography-matched controls were additionally extracted and subjected to a multiplex enzyme-linked immunosorbent assay (ELISA) assay.

“Plasma cytokines IL-1β, IL-2, and IL-6 were determined by a test developed by ARUP Laboratories (Salt Lake City, UT) and approved by the New York State Department of Health.”

Summary statistics were used to collate and analyze demographic variables and mortality statuses categorized by sex, race/ethnicity, and age. Univariate parametric tests were computed to evaluate CRS and mortality outcomes for each category. Comparisons between the mortality risks of different genotypes were conducted using multivariate logistic regressions, adjusting for sex and age.

Study findings

The present study included records from 2,589 hospitalized patients and an equal number of age, sex, and BMI-matched controls. Study participants presented a mean age of 61.2 years, an average BMI of 30.43, and comprised 53.3% male individuals.

“IL1RN rs419598, rs315952, and rs9005 genotype data were available for all patients. Biomarkers noted in the clinical electronic hospital record (EHR) for IL-1β, IL-2, and IL-6 were available for 642, 645, and 1229 subjects, respectively, whereas other plasma inflammatory markers were available for more than 2000 subjects.”

ELISA and cytokine analyses revealed that, compared to healthy control, COVID-19 patients displayed significantly elevated levels of cytokines (IL-1α, IL-5, IL-8, IL-17, IL-1β, IL-2, IL-1Ra, IL-6, tumor necrosis factor-α [TNF-α], interferon-α, and vascular endothelial growth factor [VEGF]). Alarmingly, levels of IL-6, IL-1Ra, IL-8, and IL-10 were found to be more than 10 times higher than baseline controls’ values. Inflammatory markers, including CRP, procalcitonin, D-dimer, and ferritin, were similarly heightened.

Of the included patients, 397 (15.3%) died during treatment, with age (direct), sex (male at higher risk), and BMI (direct) showing associations with COVID-19-associated mortality.

“RS-associated inflammatory biomarkers were elevated in both patients who survived and died; however, deceased patients had significantly higher levels of IL-6, CRP, procalcitonin, ferritin, and D-dimer, as well as reduced levels of complement components C3 and C4.”

Surprisingly, carriers of the IL1RN CTA-1/2 haplotype (either or two copies of the CTA haplotype) displayed substantially reduced inflammatory marker concentrations (except IL-1Ra, which was increased in these patients) compared to patients without the genotype. Encouragingly, the CTA haplotype was found to confer a 40% reduction in COVID-19-associated mortality risk in men above the age of 74. However, no associations with BMI were revealed. When evaluating each IL1RN CTA SNV individually, rs419598 C/C SNV patients exhibited substantially reduced inflammatory marker concentrations compared to their C/T or T/T counterparts.

Comparison between men and women reveals that, while most biomarker and mortality outcomes are indistinguishable across the sexes, IL1RN rs419598 C/C SNV was found to be associated with a decreased trend in mortality in men of all included age groups. In men above the age of 74, especially, this genotype was associated with an 80% decline in mortality, highlighting the role of hyperinflammation in severe COVID-19 progression.

Conclusions

The present study highlights that the IL1RN CTA haplotype, especially in combination with the rs419598 C/C genotype, substantially reduced CRS in patients (irrespective of sex) in severe COVID-19 infections and substantially reduced mortality in men.

“We show that concomitant with decreased proinflammatory cytokine production, the IL1RN CTA haplotype and rs419598 C/C SNV are associated with increased levels of its anti-inflammatory gene product IL-1Ra. Our data provide genetic evidence that activation of the inflammasome and the IL-1 pathway is proximal in the systemic cytokine inflammatory cascade. Its regulation by IL-1Ra, an endogenous anti-inflammatory protein, and potential crosstalk with IFN require further elucidation to advance the understanding and treatment of SARS-CoV-2 infection.”
Journal reference:

Attur, M., Petrilli, C., Adhikari, S., Iturrate, E., Li, X., Tuminello, S., Hu, N., Chakravarti, A., Beck, D., & Abramson, S. B. Interleukin-1 Receptor Antagonist Gene (IL1RN) Variants Modulate the Cytokine Release Syndrome and Mortality of COVID-19. The Journal of Infectious Diseases, DOI – 10.1093/infdis/jiae031, https://academic.oup.com/jid/advance-article/doi/10.1093/infdis/jiae031/7625543
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March 19, 2024
Innovative CAR T cell therapy targets two proteins to combat aggressive brain tumor growth

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In a recent report published in Nature Medicine, researchers presented the initial results of recurrent glioblastoma (rGBM) patients treated with intrathecally administered, autologous, bivalent chimeric antigen receptor (CAR) T cells targeting epidermal growth factor receptor (EGFR) and interleukin-13 receptor alpha 2 (IL13Rα2) in a phase 1 clinical trial.

Study: Intrathecal bivalent CAR T cells targeting EGFR and IL13Rα2 in recurrent glioblastoma: phase 1 trial interim results. Image Credit: Gorodenkoff/Shutterstock.com

Background

Recurrent GBM is an aggressive, treatment-resistant brain cancer with no conventional therapeutic options after chemoradiotherapy. The median overall survival (OS) is less than one year, indicating that effective treatment is an urgent unmet medical need in oncology.

Despite limited therapy options, chimeric antigen receptor T cells that target GBM-specific antigens have demonstrated tolerable safety but poor effectiveness in adults.

Tumor antigen heterogeneity, intrinsic T cell malfunction, and immunosuppressive tumor microenvironments are all examples of resistance mechanisms.

About the report

In the present report, researchers describe CART-EGFR-IL13Rα2 cell efficacy and safety in treating rGBM patients recruited in an ongoing, open-label, phase 1 trial.

The researchers included six adults presenting with multifocal and progressive wild-type glioblastoma recurring after radiotherapy, excluding those who received bevacizumab within three months before trial initiation and those with localized central nervous system tumors.

They used pre-treatment immunofluorescence analysis to detect EGFR and IL13Rα2 in brain samples. The data cutoff date was February 2, 2024, with a median follow-up of 2.5 months.

Three patients received 1 × 107 cells (dose level 1), while the others received 2.5 × 107 cells (dose level 2) 17–35 days following surgery between June 14, 2023, and January 2, 2024. The primary objectives were safety, maximum tolerated dosage, dose-limiting toxicity (DLT), and adverse events.

Secondary objectives included the proportion of treated patients, manufacturing failures, objective response rate (ORR), response length, overall survival (OS), and progression-free survival (PFS).

The team obtained brain magnetic resonance imaging (MRI) images 24 to 48 hours after CAR T therapy, four weeks later, and monthly afterward.

They collected cerebrospinal fluid (CSF) at baseline and on day one, day four, day seven, day 10, day 14, day 21, and day 28 for pharmacokinetic evaluation by quantitative polymerase chain reaction (qPCR).

They rated cytokine release syndrome (CRS) according to the American Society for Transplantation and Cellular Therapy (ASTCT) guidelines and neurotoxicity using the immune effector-associated neurotoxicity syndrome (ICANS) criteria.

They assessed treatment response using the Modified Response Assessment in Neuro-Oncology (mRANO) standards.

Results

The use of CART-EGFR-IL13Rα2 cells in cancer patients was associated with early-onset neurotoxicity, perhaps ICANS. All six patients showed reduced tumor size and enhancement, but none satisfied the ORR criteria. The researchers identified CAR T cell abundance and cytokine release in all patients and all developed neurotoxicity.

Patient 1 was diagnosed with grade 2 neurotoxicity following CAR treatment, resulting in disorientation, nausea, and aphasia. Anakinra and dexamethasone improved his neurological condition.

The researchers surgically removed a tumor nodule, revealing therapy-related alterations and uncommon glial cells. After two months, the disease progressed, for which the patient received bevacizumab and continues to remain alive with an eight-month OS.

Patient 2 had rapid tumor development and facial paralysis, which indicated grade 3 neurotoxicity. The team administered anakinra and dexamethasone as treatments. On day two, the tumor mass decreased without intervention, showing pseudo-progression. The patient developed hydrocephalus symptoms, denied shunting, and died five months after receiving CAR T cell therapy.

Patient 3, presenting with deteriorating leptomeningeal illness and decreased performance, developed grade 3 neurotoxicity and received anakinra and dexamethasone treatment.

Despite fluctuations in orientation and alertness, the patient recovered on day four, reverted to his pre-treatment neurological baseline within a week, and continues to have stable disease.

Patient 4 experienced significant neurotoxicity and received dexamethasone, anakinra, and tocilizumab, resulting in a better mental state and a restoration to the pre-treatment neurological baseline. Patient 4’s enhancement foci and periventricular nodules were decreased at treatment level 2.0 and remained stable.

Patient 5 developed multifocal tumor progression and grade 2 CAR neurotoxicity, which improved to grade 1 on the third day. He developed increased weariness, physical weakness, and anorexia. The tumor burden decreased dramatically following a doubling of dexamethasone dosage.

However, the patient returned with more lethargy, fatigue, and inadequate oral intake. On day 28, raising the steroid dosage reduced the severity of multifocal irregular enhancement.

Following a CAR T cellular injection, Patient 6 had tumor development in the left midbrain and severe right-sided hemiparesis. He experienced deteriorating aphasia and increased right-sided weakness, resulting in total hemiplegia.

Despite treatment with dexamethasone and anakinra, extensive hemiparesis remained. He got bevacizumab intravenously as part of his therapy.

The first-in-human data demonstrates the safety and bioactivity of CART-EGFR-IL13Rα2 treatment in individuals with multifocal, treatment-resistant rGBM.

The therapy decreased tumor size and enhancement but resulted in early-onset acute neurotoxicity, controllable at both dosages. The findings require confirmation with larger sample sizes and longer follow-ups.

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March 15, 2024
Man takes 217 COVID vaccines with no ill effects, shows immune boost

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In a recent case report published in The Lancet Infectious Diseases, researchers described a case of a 62-year-old male who received 217 vaccinations against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in 29 months and examined his immunological responses. They found that hyper-vaccination did not cause adverse events or significantly affect the quality of adaptive immune responses while resulting in increased T-cells and spike-specific antibodies.

Study: Adaptive immune responses are larger and functionally preserved in a hypervaccinated individual. Image Credit: Douglas Sacha / Shutterstock

Background

Booster vaccinations may potentially amplify immune responses, while persistent antigen exposure may induce immune tolerance. However, the advantages, constraints, and risks of recurrent vaccination in humans remain to be thoroughly investigated. In the present study, researchers investigated the immunological responses in an older man hyper-vaccinated against SARS-CoV-2.

The case

In this case study, a 62-year-old male from Magdeburg, Germany (referred to as HIM), engaged in deliberate hyper-vaccination against SARS-CoV-2, receiving 217 vaccinations over 29 months for personal reasons. This occurred outside a clinical study context and contrary to national recommendations. Despite an investigation by a public prosecutor for potential fraud, no criminal charges were filed. Notably, HIM’s immunological evaluation, initiated during the public prosecutor’s investigation, received active and voluntary cooperation from HIM and was ethically approved. Throughout the extensive hyper-vaccination, HIM reported no vaccine-related side effects, and routine clinical chemistry parameters displayed no abnormalities between November 2019 and October 2023. In the repeated negative SARS-CoV-2 tests, including antigen tests, polymerase chain reaction (PCR) test, and nucleocapsid serology, HIM showed no signs of past SARS-CoV-2 infection.

Starting from the 214^th vaccination, HIM’s anti-spike SARS-CoV-2 immunoglobulin G (IgG) levels were measured before and after vaccinations. The antibody peak occurred at the 214^th vaccination, and there was a slight increase after the 217^th vaccination. Additionally, HIM showed IgG4 subclass switching after the 215^th vaccination, which is uncommon in regimens with adenoviral-based vaccines as the first dose.

A total of 29 individuals who received three doses of a messenger ribonucleic acid (mRNA) vaccine formed the control group. As compared to controls, HIM exhibited mildly elevated levels of anti-spike IgM and IgA in the serum. However, in saliva samples, HIM showed detectable levels of anti-spike IgG, contrary to the control participants. HIM’s serum neutralization capacity was higher (5.4-fold for wildtype and 11.5-fold for Omicron B1.1.529 spike proteins) than the controls, indicating elevated quantities of spike-specific IgG. This observed difference was not attributed to antibody avidity as it remained comparable among the groups.

HIM showed a slightly increased number of spike-specific B-cells, with the same phenotype as seen in single-cell RNA sequencing (scRNA-seq). No significant differences were observed in the rates of somatic hypermutation or clonal expansion. CD8+ T-cells specific to the spike epitope were about six-fold more frequent in HIM, with a preference for effector memory T-cells. Further, scRNA-seq of LTD-specific T-cells showed a more differentiated phenotype and increased clonal expansion compared to controls. Flow-cytometric analysis and metabolic profiling showed no significant abnormalities in 14 protein markers.

LTD-specific CD8+ T-cells in HIM showed a proliferative capacity similar to control individuals, aligned with conserved numbers of T-cells with a phenotype like early differentiated stem cells. After epitope-specific stimulation, HIM displayed higher cytokine-positive cells, but the cytokine release per cell remained roughly equal. Cytokine analysis in the supernatant revealed the typical pattern of virus-specific CD8+ T-cells. Additionally, HIM’s CD8+ T-cells showed higher peptide sensitivity than the control group. Examination of spike-reactive CD4+ T-cells revealed a dearth of nucleocapsid-specific immunity, with similar cytokine-producing CD4+ T-cell amounts in HIM compared to the control group while retaining peptide sensitivity.

Conclusion

In conclusion, the present case report showed that hyper-vaccination against SARS-CoV-2 yielded no adverse events and elevated T-cell levels and spike-specific antibodies. Notably, the implicit quality of adaptive immune responses showed no significant effects. Although breakthrough SARS-CoV-2 infections were not observed in the individual, any causal link with the hyper-vaccination regimen remains unclear. The researchers emphasize that they do not advocate for hyper-vaccination as an approach to improve adaptive immunity.

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March 8, 2024
Schisanhenol suppresses cytokine storm and acute lung injury in mice

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Background and objectives

Cytokine storm (CS) is an acute systemic inflammatory response with limited effective interventions up to now. The treatment experience of the COVID-19 pandemic suggests great potential in the intervention of CS by herbal medicine. This study aimed to investigate whether Schisanhenol (SSH), an active component of the Chinese herbal medicine Schisandra chinensis, has the potential to interfere with CS.

Methods

The effect of SSH on nuclear factor-kappa B (NF-κB) signaling pathway activity was observed with THP-1/NF-κB cells. THP-1 and abdominal macrophages were used as cell models to observe the effect of SSH on inflammatory responses. The lipopolysaccharide-induced acute inflammatory response in mice was used to observe the effect of SSH on systemic inflammatory response and induced acute lung injury. The potential biological mechanism of SSH against inflammatory storm was explored by network pharmacology and molecular docking methods.

Results

SSH significantly inhibited NF-κB pathway activity and suppressed macrophage and systemic inflammatory responses in mice. SSH also effectively alleviated lipopolysaccharide-induced acute lung injury. The network pharmacology results showed that estimated glomerular filtration rate, matrix metalloproteinase 9, proto-oncogene tyrosine-protein kinase Src, and mammalian target of rapamycin are potential key target proteins of SSH.

Conclusions

The findings of this study demonstrate that SSH inhibited the macrophage inflammatory response and cytokine production at both the systemic and local levels in mice. Additionally, SSH effectively mitigated acute lung injury resulting from CS. Furthermore, network pharmacological analysis revealed that SSH has the ability to suppress inflammatory response through multiple mechanisms.

Source:

Journal reference:

Qi, W., et al. (2023). Schisanhenol: A Potential Drug for the Treatment of Cytokine Storm. Exploratory Research and Hypothesis in Medicine. doi.org/10.14218/ERHM.2023.00054.

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March 7, 2024
Natural killer cells lead the charge in cancer treatment innovation
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In a recent review published in the journal Nature, researchers collated available publications on natural killer (NK) cells – innate immune cells involved in recognizing and eliminating cells in distress, particularly virus-infected cells and tumors. They focus on reviewing ongoing preclinical and clinical research in the field of NK therapeutics, primarily elucidating the role of NK cells in cancer immunity. They further explore the potential for bioengineering approaches to harness NK cells via the development of genetically modified NK cells, immune checkpoint inhibitors, and cell engagers.

Review: Natural killer cell therapies. Image Credit: Numstocker / Shutterstock

What are NK cells, and why should we care?

Natural killer cells (NK cells) are innate lymphoid cells (ILCs), white blood cells that destroy infected and diseased cells, like virus-infected and cancerous cells. These cells were discovered relatively recently in 2008 and are naturally produced in the bone marrow. They can exist in populations of up to 2 x 10¹⁰ NK cells per individual, thereby representing 1% of all immune cells and 2% of all lymphocytes.

Research has revealed that in healthy humans, NK cells can be found in the liver, blood, and bone marrow, serving cytolytic and cytokine-secreting functions. Scientists have traditionally classified these immune cells into two main types based on their surface molecules – CD56 (primarily cytokine-secreting function) and CD16a (predominantly cytotoxic function). More recent RNA-based classification approaches have revealed the presence of three NK cell families:

Type 1 NK (NK1) cells correspond to the traditional CD56^dimCD16⁺ NK cells and are the most abundant in blood. They are characterized by the strong expression of CD16 (FCGR3A) and cytotoxicity effector molecules (GZMA, GZMB, and PRF1). They have recently been discovered to sometimes express genes, including SPON2, whose biological function remains to be unraveled.

Type 2 NK (NK2) cells correspond to the traditional CD56^brightCD16^– NK cells and are unique in their transcriptional signatures, chemokine profiles, cell surface markers, and their characteristic strong expression of TCF1 (a transcriptional factor). Type 3 NK (NK3) cells are the most recently discovered of these three cohorts and are characterized by CD16^dimadaptiveNKG2C^highand CD57⁺cells. The relative abundance of these cohorts has been observed to vary depending on pathophysiological conditions and anatomical localization.

NK cells have the unique properties of transitioning into an ILC1-like state, allowing them to acquire hypothesized antitumor functions. Combined with their ability to recognize cells in distress and the impressive responses of CAR T-cell therapy (modified T-cells with anti-cancer properties) and immune checkpoint inhibitors over a spectrum of malignancies, NK cells are a crucial focus of future anti-cancer therapeutics research.

What are NK cells’ anti-cancer benefits?

In addition to the aforementioned ILC1-like state, CAR T-cell therapy, and checkpoint inhibitory functions of NK cells, novel research aims at devising mechanisms by which the tumors can no longer evade T-cells and, by extension, NK cells. Unlike other T-cell populations, NK cells are not restricted by antigen-specific priming.

More applicably, NK cells are capable of recognizing cells in distress irrespective of their embryonic origin or distress trigger. NK cells are further known to produce IFNγ and similar biomolecules capable of preventing metastasis by forcing malignant cells into a state of dormancy, and FLT-3L, XCL1, and CCL5, which bolster the anti-cancer properties of dendritic cells and other lymphocytes.

“…a very important distinguishing factor between T and NK cells lies in the increase in NK cell function when tumour cells downregulate MHC-I expression on the cell surface. Loss of MHC-I expression is a common T cell immune evasion mechanism. By contrast, as NK cells express inhibitory MHC-I receptors, MHC-I loss contributes to the recognition and efficient elimination of tumour cells by NK cells. Thus, several features of NK cell biology make their use an interesting and complementary to other modalities used in oncology, including monoclonal-antibody-based therapies, cell-based therapies or a combination of both.”

Inhibitory checkpoints

Research has discovered that the activity of NK cells can be selectively switched on and off via the use of their cell-surface inhibitory receptors such as NKG2A, T cell immunoglobulin, and mucin domain-containing 3 (TIM-3), lymphocyte activation gene 3 (LAG3), and T cell immuno-receptor with Ig and immunoreceptor tyrosine-based inhibitory motif domains (TIGIT).

Preclinical and clinical trials are currently in progress to identify and test the efficacy of monoclonal antibodies in this selective activation process. For example, blocking NKG2A has been shown to unleash both NK- and T-cell-mediated antitumor responses, particularly against lung cancer. Similarly, blocking the LAG3 receptor has been shown to boost NK cell antitumor immune function; TIM-3 blocking can promote the NK-cell-mediated generalized elimination of malignant cells, while TIGIT blocking can enhance NK cell proliferation and their antitumor activities against malignant B cells.

Can NK cells be used as drug products?

A growing body of evidence suggests that NK cells can perform drug functions, especially in the field of oncological biotherapy. A number of studies are currently establishing the autologous and allogenic applications of NK cells, elucidating why, despite the apparent dearth of NK cell-based commercially available drugs, this is set to change in the near future.

“These approaches further diverge into distinct modalities, spanning from in vitro pre-activation techniques to cutting-edge genomic editing interventions. Several cancer conditions and oncological treatments, notably chemotherapy, are known to attenuate both the abundance and the operative capacity of patient’s endogenous NK cells. This depletion underscores the therapeutic rationale for adoptive NK cell transfer, a strategy to enhance the efficacy and resilience of NK cells within the TME.”

Allogenic NK cell infusions are of particular interest given their immediate bioavailability, absence of graft-versus-host disease, and robust anti-cancer potential against various malignancies. Parallel research aimed at enhancing NK cell performance is also ongoing, with ex vivo conditioning and genetic engineering presenting the most promising avenues for NK cell optimization.

Challenges to NK cells’ clinical adoption?

The present review highlights ten challenges conventional research must overcome before NK cell therapeutics receive wider medical adoption beyond current experimental procedures. These challenges can be condensed into three main aspects: 1. Improving the bioavailability of NK cells, especially at the target tumor site, 2. Enhancing the viability and cytotoxicity of NK cells, and 3. The standardization and optimization of treatment procedures using NK cells.

Conclusions

The present review explores the potential and feasibility of NK cells’ clinical applications and summarizes ongoing research on these recently discovered lymphocytes. The review reveals that despite less than two decades of research in the field, NK cells are emerging as a safe, practical, and potentially widely accessible means of clinical therapy, particularly antitumor. While challenges do exist in the adoption of NK cell therapies by mainstream medicine, studies aimed at overcoming these challenges are already underway, bringing the future of NK cell clinical interventions closer than ever.
Journal reference:

Vivier, E., Rebuffet, L., Cornen, S., Igarashi, R. Y., & Fantin, V. R. (2024). Natural killer cell therapies. Nature, 626(8000), 727-736, DOI – 10.1038/s41586-023-06945-1, https://www.nature.com/articles/s41586-023-06945-1
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February 23, 2024
Smoking, infection, and BMI found to significantly sway immune response, study shows

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In a recent study published in the journal Nature, researchers explored the factors influencing cytokine release, a critical component of the host immunological response.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic emphasized the wide variation in immunological responses between populations, with age, sex, and genetic variables all playing vital roles. However, therapy and vaccine development often disregard immunological diversity. The Milieu Intérieur research project has contributed to understanding immune homeostasis by quantitatively evaluating the impacts of age, gender, cellular composition, and genetics on immune-related gene transcript levels and those of age, gender, smoking, and cytomegalovirus (CMV) infections on leukocyte distribution in blood. Further study might help us better understand the elements that influence immune responses and how they affect clinical outcomes.

Study: Smoking changes adaptive immunity with persistent effects. Image Credit: NeydtStock / Shutterstock

About the study

In the present study, researchers investigated environmental variables associated with cytokine responsiveness to immunological activation.

The team measured the levels of several cytokines [C‐X‐C motif chemokine ligand 5 (CXCL5), colony-stimulating factor 2 (CSF2), interferon-gamma (IFNγ), interleukin-1 beta (IL-1β), IL-2, 6, 8, 10, 12p70, 13, 17, 23, and tumor necrosis factor (TNF)] after 22 hours of whole-blood stimulations with 11 immunological agonists for 1,000 Milieu Intérieur project donors and in an unstimulated (control) condition. They categorized the stimulations as microbial, viral, T-lymphocyte activated, and cytokines.

Heat maps and principal component analyses (PCA) of 13 cytokine molecules investigated in 12 immunological stimulations revealed the individual cytokines generated by every independent condition. The team performed hierarchical clustering evaluations of log mean variations in cytokine levels to identify groups corresponding to stimulation types.

The researchers compiled 136 environmental, socio-demographic, nutritional, and clinical variables from the digital case report forms and tested for their relationships with cytokines induced in every stimulation using likelihood ratio tests (LRTs) with age, experimental batch, and gender as covariates. They also investigated human leukocyte antigen (HLA) as a predictor of immune response variability, particularly in antigen-specific responses. The team investigated whether smoking-cytokine correlations continued when particular subsets of circulating immune cells were included in their models, as these cells are related to cytokine elevations. They evaluated the biological impact of smoking on cytokine production, calculating the effect sizes for the smoking variables in the linear models and assessing the influence of 326 soluble proteins in sera obtained from 400 donors.

The researchers investigated whether epigenetic pathways contribute to the impact of smoking on adaptive immune responses. They analyzed deoxyribonucleic acid (DNA) methylation at more than 850,000 CpG sites and investigated whether the levels may explain the association between smoking and cytokine levels following SEB stimulation. The study was especially well-suited to identifying response protein quantitative trait loci (pQTLs) since it tested 5,699,237 high-quality imputed single nucleotide polymorphisms (SNPs) for relationships with the cytokines elicited by each stimulation.

Results

The team identified smoking, CMV latent infection, and body mass index (BMI) as the most significant drivers of cytokine response variability. Smoking impacts innate and adaptive immune responses, with the influence on innate responses diminishing after quitting and associated with serum carcinoembryonic antigen-related cell adhesion molecule 6 (CEACAM6) levels. However, the impact on adaptive responses lasts long after smoking cessation and is associated with epigenetic memory.

The study highlighted eleven factors related to one or more cytokines in the immune stimulations, with BMI being the most prevalent. Smoking-related factors were related to interleukin-2 and interleukin-13 (adaptive immunity) in Staphylococcus aureus enterotoxin B superantigen (SEB), anti-cluster of differentiation 3 (anti-CD3) and anti-CD28 immune stimulations, and CXCL5 following Escherichia coli infections or innate immunological stimulations. The findings indicate that smoking causes inflammation and reduces immunity against bacterial infections.

Cytomegalovirus latent infection was associated with TNF, CSF2, and IFNγ cytokines secreted by adaptive immune cells. BMI-related factors were related to CXCL5 following Bacillus Calmette-Guérin (BCG) immune stimulation, and interleukin-2 following SEB stimulation demonstrated obesity dysregulation. The team found no significant association between major histocompatibility complex (MH) class II, DQ beta 1, and HLA.DBQ1.1P, and IL-6 in the control condition.

The study found 2,416 CpG locations related to smoking in the Milieu Intérieur sample, with 129 significantly associated with IL-2 in SEB stimulation. However, 11 CpGs abolished the relationship between smoking and IL-2 and IL-13. Current smokers had lower DNA methylation than non-smokers, but former smokers had an intermediate methylation level. The number of years smoked, total cigarettes smoked, and IL-2 levels in SEB stimulation were adversely linked with DNA methylation, although the number of years after smoking typically correlated positively.

Overall, the study findings identified three novel factors, i.e., smoking status, CMV latent infection, and BMI, associated with variability in cytokine secretion following immunological stimulation. These characteristics may have clinical consequences for the risk of contracting infections, cancer, or autoimmune diseases. Smokers have a heightened inflammatory response after bacterial activation, which promptly decreases after quitting. However, the impacts on adaptive immunity last for years after stopping. The link between smoking and long-lived B and T cell subsets and DNA methylation offers a potential for long-term consequences in the adaptive response.

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February 14, 2024
Ancient superseed with modern health benefits
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In a recent review published in the journal Biomedicines, researchers discuss the potential of Nigella sativa as a pharmaceutical agent.

Study: The Use of Nigella sativa in Cardiometabolic Diseases. Image Credit: M. Schuppich / Shutterstock.com

Nigella sativa: Distribution and phytoconstituents

Nigella sativa, which is commonly known as black seeds or black cumin, belongs to the Ranunculaceae family. The fruit of the Nigella sativa plant contains three to six carpels, each of which contains seeds that turn black when mature.

India is the largest producer of Nigella sativa, whereas Brazil, Colombia, Malaysia, and Canada are among the main exporters. In addition to India, Nigella sativa is also grown in Egypt, Greece, Saudi Arabia, Iran, and Pakistan.

The main bioactive components of Nigella sativa are derived from its seeds, followed by its bark and sprouts. Several primary and secondary metabolites of different chemical classes, such as alkaloids, terpenes and terpenoids, phytosterols, flavonoids, phenolic acids, and tannins, have been identified within Nigella sativa.

Nigellamines, which are the main alkaloids, and polyphenols, such as quercitrin and kaempferol, have been identified in Nigella sativa. Several fatty acids, including oleic, linoleic, and palmitic acids, have also been isolated from black cumin.

The pharmacological properties of Nigella sativa have been attributed to the presence of quinine components, particularly thymoquinone (TQ) and its derivatives, including 4‐terpineol, thymohydroquinone (THQ), p‐cymene, sesquiterpene, thymol, carvacrol, and t‐ anethol.

Medicinal use of black cumin

Black cumin has been used in traditional Chinese medicine, Arabian medicine, and Ayurveda. In Arabian medicine, Nigella sativa has been used for the treatment of asthma, bronchitis, diarrhea, indigestion, dysmenorrhea, amenorrhea, and skin infections. Comparatively, in Chinese medicine, Nigella sativa has been used as a component of formulation for the treatment of headaches.

The oil extracted from Nigella sativa seeds has antioxidant, anti‐inflammatory, immunomodulatory, and antibacterial properties. Black cumin contains tocopherols that have robust antioxidant effects, which inhibit lipid peroxidation in biological membranes.

Black cumin has been considered a natural agent that can reduce blood cholesterol. Additionally, the high levels of sterols in Nigella sativa suggest its potential use in preventing cardiovascular diseases (CVDs). Nigella sativa is also effective against diabetes, inflammatory conditions, and menopause.

Key mechanisms of Nigella sativa in alleviating cardiometabolic diseases

Several mechanisms may contribute to the protective cardiometabolic role of Nigella sativa. For example, the antioxidant properties of Nigella sativa may reduce blood pressure in patients with hypertension, whereas the anti-radical scavenging activity of TQ significantly reduces reactive oxygen species (ROS) levels.

Nigella sativa seed extracts exhibit increased catalase, superoxide dismutase, and glutathione reductase activities, which causes a significant reduction in malondialdehyde (MDA). Several studies have shown that MDA is an indicator of lipid peroxidation derived from oxidative stress. Reduced ROS levels also increase the bioavailability of nitric oxide (NO), which can significantly reduce blood pressure.

Many cardiometabolic diseases are associated with inflammation; therefore, a reduction in inflammation should have a positive effect on these diseases. Nigella sativa has been shown to inhibit inducible NO synthase, as well as reduce cytokine and pro‐inflammatory cytokine levels through inhibition of the nuclear factor κB (NF‐κB) signaling pathway. Nigella sativa has the potential to inhibit cyclooxygenase 2 (COX-2), which is an enzyme-linked with the production of inflammatory mediators, such as prostaglandins (PG).

Nigella sativa seed extracts also exhibit anti‐dyslipidemic properties. To this end, black cumin seed extract inhibits the expression of 3-hydroxy-3-methylglutaryl coenzyme A (HMG–CoA) reductase and increases the expression of low-density lipid receptors, both of which are associated with reduced cholesterol levels.

TQ acts as a peroxisome proliferator-activated receptor gamma (PPAR–γ) antagonist, which can improve insulin resistance and inhibit intestinal α‐ glucosidase, the latter of which leads to reduced glucose absorption. TQ also improves the proliferation and integrity β pancreatic cells, thereby resulting in elevated insulin secretion.

The weight reduction effect of Nigella sativa has been reported in many studies. Mechanistically, Nigella sativa induces a reduction in food intake, inhibition of intestinal glucose absorption, and increased adiponectin levels.

The cardioprotective effects of Nigella sativa seeds were demonstrated using isoproterenol‐induced myocardial injury in rats. Black cumin’s cardioprotective activity has also been associated with the mitigation of inflammation and oxidative stress.

Conclusions

Nigella sativa seeds have been used for multiple disease treatment strategies. Among the various bioactive metabolites present in black cumin, TQ is associated with most of its pharmacological properties.
Journal reference:

Derosa, G., D’Angelo, A., Maffioli, P., et al. (2024) The Use of Nigella sativa in Cardiometabolic Diseases. Biomedicines 12(2);405. doi:10.3390/biomedicines12020405
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February 14, 2024