Neutrophilic Myeloid-Derived Suppressor Cells and Severity in SARS-CoV-2 Infection.

BACKGROUND: While we strive to live with SARS-CoV-2, defining the immune response that leads to recovery rather than severe disease remains highly important. COVID-19 has been associated with inflammation and a profoundly suppressed immune response. OBJECTIVE: To study myeloid-derived suppressor cells (MDSCs), which are potent immunosuppressive cells, in SARS-CoV-2 infection. RESULTS: Patients with severe and critical COVID-19 showed higher frequencies of neutrophilic (PMN)-MDSCs than patients with moderate illness and control individuals (P = .005). Severe disease in individuals older and younger than 60 years was associated with distinct PMN-MDSC frequencies, being predominantly higher in patients of 60 years of age and younger (P = .004). However, both age groups showed comparable inflammatory markers. In our analysis for the prediction of poor outcome during hospitalization, MDSCs were not associated with increased risk of death. Still, patients older than 60 years of age (odds ratio [OR] = 5.625; P = .02) with preexisting medical conditions (OR = 2.818; P = .003) showed more severe disease and worse outcome. Among the immunological parameters, increased C-reactive protein (OR = 1.015; P = .04) and lymphopenia (OR = 5.958; P = .04) strongly identified patients with poor prognosis. CONCLUSION: PMN-MDSCs are associated with disease severity in COVID-19; however, MDSC levels do not predict increased risk of death during hospitalization.

Importance of circulating monocyte subpopulations and Fcγ receptors in immune thrombocytopenia.

Monocytes have been linked to the pathogenesis of immune thrombocytopenia (ITP) because of their role in autoantibody-mediated platelet phagocytosis. However, monocytes constitute unique populations with major differences in expression for surface Fcγ receptors (FcγRs). Thus, we evaluated monocytes in whole blood samples from patients with newly diagnosed and chronic ITP. Monocyte subpopulations were identified phenotypically by flow cytometry and defined according to the surface expression of CD14 (lipopolysaccharide receptor) and of CD16 (low-affinity Fcγ receptor III) into classical (CLM), intermediate (INTM) and nonclassical (non-CLM) monocytes. We also examined the expression of FcγRI/CD64 and FcγRIII/CD16 by monocyte subpopulations. Newly diagnosed patients showed a decrease in non-CLM, expressed as a relative percentage of total monocytes compared with controls and chronic ITP patients. Both non-CLM and INTM of newly diagnosed patients closely correlated with platelet count. These monocyte subpopulations showed significantly enhanced CD64 expression in newly diagnosed patients. On the contrary, patients with chronic ITP presented higher non-CLM in percentage than controls and concomitant lower CLM and total monocytes, in percentage and number. The expression of CD64 was increased by all monocyte subpopulations, CLM, INTM, and non-CLM in chronic patients. In conclusion, differences in monocyte subpopulations, together with enhanced expression of FcγRI/CD64 are evident in patients with ITP.

Clinical Significance of Evaluation of Monocytic Receptors in Patients with Hepatitis C Virus Infection.

Monocytes in hepatitis C virus (HCV) infection play a critical role in chronic liver inflammation and fibrosis. We studied circulating monocytes and monocyte receptors in patients with HCV infection who were naive to treatment and those who received direct acting antiviral therapy and achieved sustained virological response. CD64+ CCR2+ (M1-like) and CD206+ CD163+ CX3CR1+ (M2-like) monocyte numbers and receptor expression were evaluated by flow cytometry. Higher expression of the monocyte chemokine receptor CCR2 predicted the severity of liver fibrosis, independent of successful treatment and viral clearance (R2 = 0.235, p = 0.002), whereas monocyte CX3CR1 expression was lower in both treated and untreated patients compared with controls (p = 0.011). The expression of the scavenger receptor CD163 was lower in patients with successful treatment (p = 0.005), supporting its role as a marker of treatment response. CD64+ CCR2+ (M1-like) and CD206+ CD163+ CX3CR1+ (M2-like) monocyte numbers were not altered with fibrosis progression or treatment response. Our findings reflect the diverse functions of monocytes in liver inflammation, fibrosis, and therapy. However, HCV clearance did not lead to complete monocyte reconstitution. Targeting monocytes and their chemokine receptors bears therapeutic potential to reduce liver fibrosis and improve disease outcome.

Serum IFNγ-induced protein 10 (IP10/CXCL10): association with asthma exacerbations and severity in children.

OBJECTIVE: Asthma is a chronic disorder of the airways, in which chemokines coordinate airway inflammation and determine its severity. We aimed to study the chemokine interferonγ-inducible protein 10 kDa (IP10/CXCL10), a member of the CXC receptor 3 (CXCR3) ligand family, at the protein level in the serum of children, to evaluate the association between CXCL10 and exacerbations of childhood asthma. METHODS: Patients experiencing an asthma exacerbation (42 patients) and stable patients (43 patients) were investigated for serum CXCL10 levels. RESULTS: Patients with an asthma exacerbation expressed significantly higher CXCL10 levels in the serum than stable patients (p < 0.001). Additionally, CXCL10 values were elevated in severe asthma compared with moderate and mild disease (p < 0.001). In patients experiencing asthma exacerbations, higher values of CXCL10 were observed in atopic patients compared with non-atopic patients (p = 0.027) and in uncontrolled and partly controlled patients compared with controlled patients (p = 0.046). CONCLUSIONS: CXCL10 is proposed as an inflammatory serum marker for asthma exacerbations and worsening asthma symptoms. The levels of CXCL10 are representative of the clinical severity of asthma.

Donor cell microchimerism in kidney transplantation: Implications for graft function.

Given the uncertainty regarding the relationship between donor cells at microchimeric levels and its influence on graft function and clinical outcome, we explored the extent and importance of donor microchimerism in kidney transplantation. Twenty patients with chronic kidney disease who had received allografts from living donors were studied. We examined peripheral whole blood samples from the recipients one month after the transplant, applying mitochondrial DNA variant-specific polymerase chain reaction (PCR) to identify and quantify donor cells in relation to allograft function and survival during three years of follow-up. Higher quantities of donor-derived cell microchimerism in the peripheral blood correlated with better graft function in the early postoperative period at 1 month (R2  = .536, p = .001) and predicted improved graft function 1 year following the transplant (R2  = .430, p = .008). Furthermore, early post-transplant quantities of donor cell microchimerism were an important predictor of improved kidney function 3 years after transplantation (R2  = .397, p = .021). However, donor cell microchimerism failed to predict patient and graft survival after 3 years (odds ratio = 0.536, p = .860). Our findings suggest that donor cell microchimerism plays an immunoregulatory role in kidney transplantation and contributes to donor-specific immune hypo-responsiveness and graft acceptance.

Neutrophil and monocyte receptor expression in patients with sepsis: implications for diagnosis and prognosis of sepsis.

Understanding the complex immune responses in sepsis is crucial to provide insight into the clinical syndrome. We evaluated the changes in the surface receptors of the cells of innate immunity, neutrophils and monocytes, in patients with sepsis. Since sepsis remains a clinical challenge, we aimed to assess the significance of altered receptor expression in diagnosis and prognosis. Critically ill patients with sepsis (n=31) were investigated for the expression of receptors for IgG heavy chain CD64 and CD16 on neutrophils and CD64 and the lipopolysaccharide receptor CD14 on monocytes by flow cytometry and compared to 23 patients with no sepsis. Patients with sepsis had increased expression of neutrophil CD64. Neutrophil CD64 was specific for discriminating patients with sepsis but showed weak sensitivity. When integrated in a scoring system, neutrophil CD64 in combination with C-reactive protein (CRP) and SOFA score showed a diagnostic accuracy of 0.93 for sepsis and significantly predicted increased mortality risk. While neutrophil CD16 did not discriminate for sepsis, decreased expression was associated with increased mortality risk. In contrast, monocyte CD64 and CD14 expression was unaltered in sepsis and was not associated with mortality risk. Our study demonstrates that unlike monocytes, neutrophil receptor expression is altered in patients with sepsis receiving intensive care. It is promising to apply a combination approach to diagnose sepsis especially in time-limited conditions.

Chimerism in pediatric hematopoietic stem cell transplantation and its correlation with the clinical outcome.

Hematopoietic stem cell transplantation (HSCT) is the only hope to cure many inherited and acquired hematological disorders in children. Monitoring of chimerism helps to predict the post-transplantation events, with the intention to enhance the long-term disease free survival (DFS). The study aimed to investigate the importance of early chimerism detection to predict the clinical outcome following HSCT. The study included nine recipients (six ß-thalassemia and three severe aplastic anemia patients) and their 10/10 HLA identical sibling donors. Chimerism detection was performed by analysis of short tandem repeat (STR) polymerase chain reaction (PCR) for detection and quantification of the relative amounts of donor and recipient cells present on day +28. Peripheral blood (PB) was the main stem cell source for HSC transplantation. Disease free survival (DFS) was 71.4% while overall survival was 85.7% for PBSC transplants at the median follow up period of 4years. The early detection of chimerism by PCR-STR analysis for children with ß-thalassemia and aplastic anemia correlated with the outcome of HSCT in 8 (88.8%) patients. Complete chimerism was associated with disease-free survival while mixed chimerism and autologous patterns were associated with poor prognosis. In conclusion, early chimerism testing is clinically important in prediction of outcome after allogeneic HSC transplantation.

Serum Amyloid A Type 1 Gene Polymorphism in Egyptian Children with Familial Mediterranean Fever.

BACKGROUND: Since spontaneous inflammation is an important contributor to familial Mediterranean fever (FMF), genetic variants mediating inflammation are of interest. We investigated gene variants in the acute-phase serum amyloid A type 1 (SAA1), a sensitive marker of inflammatory activity, and their association with susceptibility and severity of FMF. METHODS: The genotypes of 2 single-nucleotide polymorphisms within exon 3 of SAA1 (2995C/T and 3010C/T) were determined in 105 Egyptian children with FMF and in 125 controls by polymerase chain reaction-restriction fragment length polymorphism. Genotyping of the causative MEFV mutations was performed by reverse hybridization. RESULTS: The M694I mutation was the most frequent allele (42.8%), followed by V726A (18.6%), M680I (17.1%), E148Q (11.9%) and M694V (9.0%). The frequency of the SAA1 α, ß and x03B3; alleles was not significantly different between FMF patients and controls. The genotype frequency of SAA1 α/α was higher in patients than in healthy subjects (21.0 vs. 14.4%) although it did not reach statistical significance. The clinical manifestations including age at disease onset, number of FMF attacks, colchicine dose and severity score were not related to genotypes of SAA1. However, M694V mutation and female gender were significantly associated with severity. CONCLUSION: The genetic polymorphism of SAA1 is not associated with susceptibility and severity of FMF in Egyptian children.

Polymorphisms of Immunoglobulin G Fc Receptors in Pediatric Guillain-Barré Syndrome.

Introduction Guillain-Barré syndrome (GBS) is an autoimmune peripheral neuropathy characterized by demyelination and axonal damage. Biallelic functional polymorphisms in the immunoglobulin G Fc receptors (FcγR)-FcγRIIA: H131/R131, FcγRIIIA: V158/F158, and FcγRIIIB: NA1/NA2 affect the affinity of the IgG-FcγR interaction, therefore, diseases such as GBS in which this interaction plays a critical role might be influenced by the polymorphisms. Methods We evaluated the role of FcγR polymorphisms in susceptibility to GBS in Egyptian pediatric patients and the association of the variant alleles with neurophysiological types, severity, and outcome of the disease. A total of 50 patients with GBS and 50 controls were examined for FcγR polymorphisms by allele-specific polymerase chain reaction. Results FcγRIIA H131 allele (p = < 0.0001; odds ratio [OR] = 4.78; 95% confidence interval [CI], 2.62-8.70) and FcγRIIA H/H131 genotype (p = < 0.0001 ; OR = 10.56; 95% CI, 3.59-31.06) were significantly increased in GBS patients while FcγRIIIA and FcγRIIIB allelic distributions were similar among patients and controls. The FcγR genotypes showed no association with neurophysiological types of GBS, severity or outcome of the disease. Conclusions These findings reflect that FcγRIIA H131 allele may represent a risk marker for susceptibility to GBS.

Down-regulation of expression of retinoid acid-related orphan receptor C (RORC) in systemic lupus erythematosus.

CONTEXT: Retinoic acid-related orphan receptor C (RORC), the key factor orchestrating the transcription of genes encoding interleukin 17, plays a critical role in the regulation of inflammatory responses. OBJECTIVE: The objective of this study was to analyze the expression of RORC in the peripheral blood of patients with systemic lupus erythematosus (SLE) for a better understanding of the pathogenesis of SLE especially in relation to disease activity and clinical and biochemical findings. METHODS: The study included 24 patients with SLE and a control group of 18 healthy gender- and age-matched individuals. Evaluation of the level of expression of RORC mRNA was performed by real-time polymerase chain reaction. RESULTS: The results showed that patients with SLE had lower RORC gene expression levels compared with healthy subjects that were not correlated with disease activity. The down-regulation of RORC was significantly lower in patients with lupus nephritis in remission than active lupus nephritis and nonrenal patients. CONCLUSIONS: The findings suggest that RORC plays a significant role in the dysregulated immune response associated with SLE. Deciphering the intricate regulatory network and the target genes of RORC will help unravel new specific treatments for SLE.

Passenger lymphocyte syndrome in ABO and Rhesus D minor mismatched liver and kidney transplantation: A prospective analysis.

The increasing demand for solid organs has necessitated the use of ABO and Rhesus (Rh) D minor mismatched transplants. The passenger lymphocyte syndrome (PLS) occurs when donor lymphocytes produce antibodies that react with host red blood cell (RBC) antigens and result in hemolysis. Our aim was to evaluate prospectively the role of PLS in post transplant anemia and hemolysis in ABO and RhD minor mismatched recipients of liver and kidney grafts and to study the association of PLS with donor lymphocyte microchimerism. We examined 11 liver and 10 kidney recipients at Day +15 for anemia, markers of hemolysis, direct antiglobulin test and eluates, and serum RBC antibodies. Microchimerism was determined in peripheral blood lymphocytes by genotyping of simple sequence length polymorphisms encoding short tandem repeats. Immune hemolytic anemia and anti-recipient RBC antibodies were observed in 2 out of 11 liver (18.2%) and 2 out of 10 kidney (20%) transplants. RBC antibody specificity reflected the donor to recipient transplant, with anti-blood group B antibodies identified in 2 cases of O to B and 1 case of A to AB transplants while anti-D antibodies were detected in 1 case of RhD-negative to RhD-positive transplant. Donor microchimerism was found in only 1 patient. In conclusion, passenger lymphocyte mediated hemolysis is frequent in minor mismatched liver and kidney transplantation. Recognizing PLS as a potential cause of post transplant anemia may allow for early diagnosis and management to decrease the morbidity and mortality in some patients.

Susceptibility and progression of end stage renal disease are not associated with angiotensin II type 1 receptor gene polymorphism.

CONTEXT: The role of the angiotensin II type 1 receptor (AT1R) gene polymorphism, A1166C, has been shown to be associated with end stage renal disease (ESRD) and its progression. There is also some evidence that HLA class II alleles are associated with ESRD independent of other factors. OBJECTIVE: To examine the association between AT1R gene polymorphism in the susceptibility and progression to ESRD in patients with chronic renal failure and to investigate if the AT1R genotypes and HLA-DR alleles predict the time to ESRD. MATERIALS AND METHODS: Genotyping was performed in 50 ESRD patients and 44 control subjects for the AT1R A1166C gene polymorphism using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). ESRD patients were examined for HLA-DRB1 alleles according to a reverse hybridization line probe assay. RESULTS: Allele and genotype frequencies of the AT1R polymorphism did not differ significantly between ESRD patients and controls. Furthermore, there was no association between the AT1R gene polymorphism or HLA-DRB1 alleles with the time to the occurrence of end stage failure. DISCUSSION AND CONCLUSION: We concluded that the AT1R genotype does not contribute to the genetic susceptibility of ESRD and is not associated with progression of chronic kidney failure to ESRD.

Increased DNA damage in hepatitis C virus-related hepatocellular carcinoma.

One consequence of hepatitis C virus (HCV) infection is an elevated cancer risk. During chronic viral infection, deoxyribonucleic acid (DNA) damage is being induced by reactive oxygen and nitrogen species, which may play a pathogenic role in HCV-induced carcinogenesis. The study investigated DNA damage in peripheral blood lymphocytes from patients with hepatocellular carcinoma (HCC) and those with HCV infection with and without associated cirrhosis and normal controls. As a measure for genomic damage, the comet assay (single cell gel electrophoresis) was applied, which detects single- and double-strand breaks and alkali-labile sites through electrophoretic mobility of the resulting fragments. The levels of DNA damage were significantly higher in HCC and HCV-associated cirrhosis compared to HCV without cirrhosis and the control group. Patients presenting with DNA damage more than mean+two standard deviation of the controls had a 3.6-fold risk of having HCC more than those with undamaged DNA. HCV disease progression was the only discriminator predicting the extent of DNA damage. The accumulation of DNA damage is important in HCC evolution. DNA damage indicating intracellular oxidative and nitrative stress may lead to mutagenesis and consequently malignant transformation, which emphasizes the need to optimize the therapy for reducing the degree of genomic damage.

Evaluation of cytokines in follicular fluid and their effect on fertilization and pregnancy outcome.

Cytokines in follicular fluid (FF) are important for reproduction as they modulate oocyte maturation and ovulation which influence subsequent fertilization, development of early embryo and potential for implantation. We evaluated FF cytokines in women who underwent intracytoplasmic sperm injection (ICSI) and their association with fertilized oocytes, embryo quality and pregnancy outcome. FF belonging to 38 patients including 18 polycystic ovary (PCO) and 20 male/unexplained infertility patients were investigated for granulocyte colony stimulating factor (G-CSF), regulated upon activation normal T cell expressed and presumably secreted (RANTES), tumour necrosis factor (TNFα), interferon gamma (IFNγ) and interleukins (IL-4 and IL-2) by bead-based sandwich immunoassay. Our findings revealed that on the day of oocyte retrieval, G-CSF was positively correlated with the number of fertilized oocytes, while TNFα detection was associated with reduced number of fertilized oocytes. Only G-CSF showed significant positive effect to the pregnancy outcome although the cytokines studied were not associated with embryo quality. PCO as the cause of infertility did not show an association with cytokines in FF. The functions of cytokines in reproduction are likely to be complex, and cytokine evaluation may offer insight to the understanding of the mechanisms leading to success or failure of assisted reproduction.

Asynchronous DNA replication and aneuploidy in lymphocytes of hepatocellular carcinoma patients.

Using fluorescence in situ hybridization (FISH) analysis, we examined the replication mode of the centromere region (homologous counterpart) and the aneuploidy level of chromosome 17 in the interphase nuclei of phytohaemagglutinin (PHA)-stimulated peripheral blood lymphocytes from (1) patients with hepatocellular carcinoma (HCC); (2) patients with liver cirrhosis (LC) due to hepatitis C viral infection who are individuals at a higher increased risk for HCC; and (3) healthy control participants. We also compared the allelic-replication asynchrony and aneuploidy frequencies with serum alpha-fetoprotein (AFP) levels. We found a significant increase in centromeric replication asynchrony accompanied by a high frequency of aneuploidy in lymphocytes of HCC patients compared with those of LC patients and healthy control participants. These changes are similar to those previously observed in other types of malignancy (hematological, ovarian, prostate, and breast cancer). The cytogenetic alterations of aneuploidy and strong asynchronous replication displayed in the lymphocytes of HCC patients arose from malignancy, as they were associated neither with an increased risk for cancer nor with an infection. The cytogenetic cancer-associated markers observed in patients' lymphocytes appeared to be superior to serum AFP, the marker currently used for HCC. Thus, the cytogenetic cancer-associated markers may be potentially useful in noninvasive cancer detection.