Peripheral Blood Mononuclear Cell Reactivity Research Articles

Differential peripheral blood mononuclear cell reactivity against SARS-CoV-2 proteins in naïve and previously infected subjects following COVID-19 vaccination.

The decline in vaccine efficacy and the risk of reinfection by SARS-CoV-2 make new studies important to better characterize the immune response against the virus and its components. Here, we investigated the pattern of activation of T-cells and the expression of inflammatory factors by PBMCs obtained from naïve and previously infected subjects following COVID-19 vaccination, after PBMCs stimulation with S1, RBD, and N-RBD SARS-CoV-2 proteins. PBMCs showed low levels of ACE2 and TMPRSS2 transcripts, which were not modulated by the exposure of these cells to SARS-CoV-2 proteins. Compared to S1 and RBD, N-RBD stimulation showed a greater ability to stimulate T-cell reactivity, according to CD25 and CD69 markers. Interestingly, T-cell reactivity was more pronounced in vaccinated subjects with prior SARS-CoV-2 infection than in vaccinated donors who never had been diagnosed with COVID-19. Finally, N-RBD stimulation promoted greater expression of IL-6 and IFN-γ in PBMCs, which reinforces the greater immunogenic potential of this protein in the vaccinated subjects. These data suggest that PBMCs from previously infected and vaccinated subjects are more reactive than those derived from just vaccinated donors. Moreover, the N-RBD together viral proteins showed a greater stimulatory capacity than S1 and RBD viral proteins.

A template for physical resilience research in older adults: Methods of the PRIME-KNEE study.

Older adults with similar health conditions often experience widely divergent outcomes following health stressors. Variable recovery after a health stressor may be due in part to differences in biological mechanisms at the molecular, cellular, or system level, that are elicited in response to stressors. We describe the PRIME-KNEE study as an example of ongoing research to validate provocative clinical tests and biomarkers that predict resilience to specific health stressors. PRIME-KNEE is an ongoing, prospective cohort study that will enroll 250 adults ≥60 years undergoing total knee arthroplasty. Data are collected at baseline (pre-surgery), during surgery, daily for 7 days after surgery, and at 1, 2, 4, and 6months post-surgery. Provocative tests include a cognition-motor dual-task walking test, cerebrovascular reactivity assessed by functional near-infrared spectroscopy, peripheral blood mononuclear cell reactivity ex vivo to lipopolysaccharide toxin and influenza vaccine, and heart rate variability during surgery. Cognitive, psychological, and physical performance batteries are collected at baseline to estimate prestressor reserve. Demographics, medications, comorbidities, and stressor characteristics are abstracted from the electronic medical record and via participant interview. Blood-based biomarkers are collected at baseline and postoperative day 1. Repeated measures after surgery include items from a delirium assessment tool and pain scales administered daily by telephone for 7 days and cognitive change index (participant and informant), lower extremity activities of daily living, pain scales, and step counts assessed by Garmin actigraphy at 1, 2, 4, and 6months after surgery. Statistical models use these measures to characterize resilience phenotypes and evaluate prestressor clinical indicators associated with poststressor resilience. If PRIME-KNEE validates feasible clinical tests and biomarkers that predict recovery trajectories in older surgical patients, these tools may inform surgical decision-making, guide pre-habilitation efforts, and elucidate mechanisms underlying resilience. This study design could motivate future geriatric research on resilience.

High-fat meal increases peripheral blood mononuclear cell pro-inflammatory cytokine expression in African-American women.

African-American (AA) women have elevated predominance of inflammatory diseases concurrent with local inflammation resulting in compromised metabolic function. The purpose of the study was 2-fold: 1) to examine the gene and protein expression of pro- and anti-inflammatory cytokine secretion by peripheral blood mononuclear cells (PBMC) obtained from AA and Caucasian-American (CA) women in response to an acute high-fat meal; and 2) to explore the influence of race (AA vs. CA) on PBMC reactivity. Ten AA and 11 CA women consumed a high-fat meal with baseline and 4h postprandial venous blood draws. PBMCs were incubated for 3 h then messenger RNA expression and supernatant protein concentration was used to examine inflammatory profiles. All women had a postprandial increase in interleukin (IL)-8 gene expression, IL-8 protein concentration, and tumor necrosis factor alpha (TNF-α) protein concentration (P< 0.05). AA women had a postprandial increase in IL-6, IL-8, and TNF-α protein concentration (P< 0.05). AA women had higher postprandial IL-1β protein concentration and IL-8 gene expression compared with CA women (P< 0.05). Our data uncovers the specific impact of race and time on pro-inflammatory PBMC (IL-1β, IL-6, IL-8, and TNF-α) expression profiles in response to an acute high-fat meal challenge. Novelty: African Americans have higher predominance of inflammatory disease. We explored the potential race impact on peripheral blood mononuclear cell reactivity in response to a meal. A pro-inflammatory response to an acute high-fat meal with race impact was observed possibly contributing to health disparities impacting African-American women.

1679-P: Islet Autoimmunity in Adults with Impaired Glucose Tolerance (IGT) and Newly Diagnosed, Treatment-Naïve Type 2 Diabetes (NDX T2D) in the Restoring Insulin Secretion (RISE) Study

In obesity, immune cells establish a systemic inflammatory state associated with development of insulin resistance and T2D. Previous studies have shown that peripheral blood mononuclear cell (PBMC) responses to islet proteins (IPs) in adults with established T2D on glucose-lowering therapies identify those patients with more severe β-cell dysfunction. Moreover, attenuation of PBMC responses to IPs in adults with established T2D improves β-cell function. However, the presence of IP reactive PBMCs, glutamic acid decarboxylase autoantibodies (GADA), and their relationship to insulin sensitivity and β-cell function has never been evaluated in IGT or Ndx T2D. We measured at baseline and after 12 months of RISE medication interventions (12 months with either metformin or liraglutide plus metformin or for 3 months with insulin glargine followed by 9 months of metformin), PBMC responses to IPs, GADA, insulin sensitivity, and β-cell function in 258 RISE participants (mean ± SD) 53.3 ± 9.2 years of age, BMI 35.1 ± 5.4 kg/m2, with IGT (n=180) or Ndx T2D (&amp;lt;12 months duration, n=78). At baseline, IP reactive PBMCs were present in 46% of participants with IGT and 40% of participants with Ndx T2D. However, 97% of IGT and 98% of Ndx T2D were GADA negative. There were no significant relationships between the presence of PBMC reactivity to IPs, GADA positivity and insulin sensitivity or β-cell function. RISE treatments did not alter PBMC responses to IPs, GADA, or the relationship between IP reactive PBMCs or GADA with glycemic responses. In summary, IP reactive PBMCs are present in a large percentage of adults with very early dysglycemia, However, the presence of IP reactive PBMCs did not correlate with insulin sensitivity or early β-cell dysfunction. Further studies are needed to unravel the pathogenesis of IP reactive PBMCs and GADA and their effects on β-cell function during progression of T2D. Disclosure B. Brooks-Worrell: None. A.H. Tjaden: None. S. Edelstein: None. B.M. Palomino: None. K. Utzschneider: Other Relationship; Self; Medtronic. S.A. Arslanian: Research Support; Self; National Institutes of Health. Other Relationship; Self; AstraZeneca, Eli Lilly and Company, Novo Nordisk Inc. K.J. Mather: Research Support; Self; Abbott, Merck &amp; Co., Inc., Novo Nordisk Inc., Sanofi. T.A. Buchanan: None. K.J. Nadeau: None. K.M. Atkinson: None. E. Barengolts: None. S.E. Kahn: Advisory Panel; Self; Boehringer Ingelheim International GmbH, Eli Lilly and Company, Intarcia Therapeutics, Janssen Scientific Affairs, LLC., Merck &amp; Co., Inc., Novo Nordisk A/S, Pfizer Inc. J.P. Palmer: None. Funding National Institutes of Health (R01DK107560)

Administering 25-hydroxyvitamin D3 in vitamin D-deficient young type 1A diabetic patients reduces reactivity against islet autoantigens

We investigated whether improving 25-hydroxyvitamin D status in young type 1A diabetic patients reduces reactivity of peripheral blood mononuclear cells against islet autoantigens and associates with beta-cell functional changes. Eight patients with 25-hydroxyvitamin D deficiency (<20ng/ml), out of 15 consecutive young type 1A diabetic subjects received 25-hydroxyvitamin D3 to achieve and maintain levels above 50ng/ml for up to one year. Peripheral blood mononuclear cell reactivity (Interferon-γ spots) against beta-cell autoantigens (glutamic acid decarboxylase 65-kD isoform, proinsulin and tyrosine phosphatase-like protein IA-2) and C-peptide during mixed meal were assessed before and after 25-hydroxyvitamin D3 replenishment. Target 25-hydroxyvitamin D blood levels were safely reached and maintained. Peripheral blood mononuclear cell reactivity against glutamic acid decarboxylase 65-kD isoform (3.8±4.0 vs. 45±16) and proinsulin (3.5±3.2 vs. 75±51) decreased significantly (p<0.001 and p<0.02) upon 25-hydroxyvitamin D3 replenishment, which was correlated with 25-hydroxyvitamin D concentrations. C-peptide values remained stable after one year of treatment. Safely restored and maintained 25-hydroxyvitamin D levels associated with reduced peripheral blood mononuclear cell reactivity against beta-cell autoantigens with no significant decrease of beta-cell function in this cohort of patients.

Peripheral Blood Mononuclear Cell Reactivity to LPS Is Related to Granuloma Severity in Sarcoidosis

PURPOSE: Granuloma formation is a common inflammatory condition in the lungs of patients with sarcoidosis. The underlying mechanism is unknown but several studies suggest that fungi in the environment may be related to the risk of disease.

Cholecalciferol Plus Calcium Suppresses Abnormal PBMC Reactivity in Patients with Multiple Sclerosis

The active metabolite of vitamin D, 1,25-dihydroxyvitamin D [1,25(OH)(2)D], is a potent modulator of immune cells in vitro. Our objective was to determine whether the sun-dependent nutrient, cholecalciferol, can alter disease-associated cellular immune abnormalities in patients with multiple sclerosis (MS). This was an open-label, 12-month, randomized controlled trial. Patients with MS were recruited from the MS Clinic at St. Michael's Hospital, Toronto. Forty-nine patients were matched (for age, sex, disease duration, disease-modifying drug, and disability) and enrolled (treated n = 25; control n = 24). Four patients were lost to follow-up (n = 2 from each group). Treated patients received increasing doses of cholecalciferol (4,000-40,000 IU/d) plus calcium (1200 mg/d), followed by equilibration to a moderate, physiological intake (10,000 IU/d). Control patients did not receive supplements. At enrollment and at 12 months, peripheral blood mononuclear cell (PBMC) proliferative responses to disease-associated, MS-relevant, and control antigens were measured, along with selected serum biochemical markers. At 12 months, mean serum 25-hydroxyvitamin D [25(OH)D] concentrations were 83 ± 35 nmol/liter and 179 ± 76 nmol/liter in control and treated participants, respectively (paired t, P < 0.001). Serum 1,25(OH)(2)D did not differ between baseline and 1 yr. In treated patients, 12-month PBMC proliferative responses to neuron antigens myelin basic protein and exon-2 were suppressed (P = 0.002). In controls, there were no significant changes in disease-associated PBMC responsiveness. There were no significant differences between groups in levels of selected biomarkers. MS-associated, abnormal T cell reactivities were suppressed in vivo by cholecalciferol at serum 25(OH)D concentrations higher than 100 nmol/liter.

Reactivation of type 1 diabetes in patients receiving human fetal pancreatic tissue transplants without immunosuppression.

Type 1 diabetes is a cell-mediated autoimmune disease. Successful transplantation of human fetal pancreatic tissue into type 1 diabetic patients must address both autoimmunity and allograft rejection. We investigated whether humoral and cellular responses to islet antigens could be demonstrated in the peripheral blood of type 1 diabetic subjects receiving human fetal pancreatic tissue transplants. We investigated peripheral blood mononuclear cell (PBMC) responses, using cellular immunoblotting, and autoantibody responses to islet proteins, before transplant and at 3-month intervals after transplant, of nine long-term type 1 diabetes patients (mean disease duration of 21 years) receiving human fetal pancreatic tissue subcutaneously into the abdominal wall without immunosuppression. Before transplant, all nine subjects were islet cell autoantibody (ICA)-negative and seven out of nine subjects were glutamic acid decarboxylase autoantibody (GADAb)-positive. After transplant, all subjects became ICA(+) and the two patients who were GADAb(-) before transplant became GADAb(+) after transplant. Maximum PBMC reactivity to separated human fetal pancreatic proteins was observed in four patients 3 months after transplant, in one patient at 6 months, in two patients at 9 months, and in one patient at 12 months after transplant. One subject, who had PBMC reactivity to multiple islet proteins before transplant, continued to respond to multiple islet proteins throughout the study. We conclude that the development in the peripheral blood of ICA, GADAb, and PBMC reactivity to human fetal pancreatic proteins in the trans plant recipients is most consistent with reactivation of the type 1 diabetes disease process.

Peripheral blood mononuclear cell responses from type 1 diabetic patients and subjects at-risk for type 1 diabetes to human fetal pancreatic tissue proteins.

Fetal pancreatic tissue has been suggested to be less immunogenic than adult islets. Thus, transplantation of human fetal pancreatic tissue as treatment for type 1 diabetes has been gaining interest. To investigate this question, we tested the peripheral blood mononuclear cell (PBMC) responses from different subject populations to human adult islet proteins (AIP) versus human fetal pancreatic proteins (FPP). PBMC responses to FPP and AIP from normal controls (n=14), newly diagnosed type 1 diabetes patients (n=5), long-term type 1 diabetes patients (n=9), and subjects at-risk for development of type 1 diabetes (n=3) were studied. We observed that normal controls demonstrated PBMC reactivity to 0-3 molecular weight regions (mwr) for both the AIP (mean+/-SD, 0.8+/-1.1) and the FPP (0.6+/-0.7). In contrast, newly diagnosed type 1 diabetic patients (<1 year) demonstrated PBMC responses to 9-16 mwr for the AIP (12.8+/-2.5) and 0-14 mwr for the FPP (6.8+/-5.0). The PBMCs from long-term type 1 diabetes patients (> 3 years) were responsive to 2-11 mwr for AIP (6.0+/-2.8) and 0-11 mwr for FPP (4.9+/-4.0). Three nondiabetic ICA positive subjects at-risk for development of type 1 diabetes demonstrated positive PBMC reactivity to 9-18 mwr for the AIP (12.7+/-3.9) and 4-18 mwr for the FPP (10.0+/-5.9). We conclude that human fetal pancreatic proteins are not significantly less stimulatory than human adult islet proteins to PBMCs of subjects with or at risk for type 1 diabetes.

Reactivation of type 1 diabetes in patients receiving human fetal pancreatic tissue transplants without immunosuppression.

Insulin-dependent (type 1) diabetes is a cell-mediated autoimmune disease. Successful transplantation of human fetal pancreatic tissue into type 1 diabetic patients must address both autoimmunity and allograft rejection. We investigated whether humoral and cellular responses to islet antigens could be demonstrated in the peripheral blood of type 1 diabetic subjects receiving human fetal pancreatic tissue transplants. We investigated peripheral blood mononuclear cell (PBMC) responses, using cellular immunoblotting, and autoantibody responses to islet proteins, before transplantation and at 3-month intervals after transplantation. Our study population included nine long-term type 1 diabetes patients (mean disease duration of 21 years) receiving human fetal pancreatic tissue subcutaneously into the abdominal wall without immunosuppression. Before transplantation, all nine subjects tested negative for islet cell autoantibody (ICA), and seven of nine subjects tested positive for glutamic acid decarboxylase autoantibody (GADAb). After transplantation, all subjects became ICA(+), and the two patients who were GADAb(-) before transplantation, became GADAb(+) after transplantation. Maximum PBMC reactivity to separated human fetal pancreatic proteins was observed in four patients at 3 months, in one patient at 6 months, in two patients at 9 months, and in one patient at 12 months after transplantation. One subject, who had PBMC reactivity to multiple islet proteins before transplantation, continued to respond to multiple islet proteins throughout the study. We conclude that the development in the peripheral blood of ICA, GADAb, and PBMC reactivity to human fetal pancreatic proteins in the transplant recipients is most consistent with reactivation of the type 1 diabetes disease process.

Influenza vaccination in a healthy geriatric population: preferential induction of antibodies specific for the H3N2 influenza strain despite equal T cell responsiveness to all vaccine strains

Cellular as well as humoral immune reactivity were studied in healthy young (<30 years; n = 12) and older (> 65 years; n = 12) individuals before as well as 1 month after immunization with a trivalent whole virus influenza vaccine. Before vaccination, peripheral blood mononuclear cell proliferation in response to in vitro stimulation with each of the virus strains was low in both groups. No antibodies against either the H1N1 or the B strain were found in most individuals, while 91% of the young and 75% of the elderly persons had low but protective antibody titres to the H3N2 strain. Vaccination led to a significant enhancement of peripheral blood mononuclear cell reactivity to all three influenza strains in both age groups. However, there was a significant difference in the humoral immune response between the groups. While there was a vigrous antibody response to all three vaccine strains among young persons, protective titres against the H1N1 and the B strains were only just reached in the old. In contrast, antibody production to the H3N2 strain was most abundant in the majority of elderly individuals, leading to significantly higher titres in the old than in the young group. In conclusion, the results demonstrate the preferential induction of antibodies to one particular influenza strain despite equal T cell recruitment to all vaccine strains in healthy aged individuals after immunization with a trivalent influenza vaccine. Our findings underline the complexity of immunological alterations to be expected after vaccination in healthy elderlies.

Angiotensin II Suppkksion of Human Mononuclear Cell React I VI TY is Associated with Enhanced OKT8+Lymphocyie Thymidine Incorporation

Recent evidence suggests that angiotensin II may participate in the regulation of inflammation. We previously reported that angiotensin II inhibits human peripheral blood mononuclear cell reactivity and acts directly on lymphocytes. These observations are again confirmed. In addition, purified OKT8+ but not OKT4+ T cell suspensions stimulated with phytohemagglutinin revealed increased thymidine incorporation when simultaneously cultured for 48 hours with angiotensin II. These findings suggest that angiotensin II may inhibit mononuclear cell thymidine uptake through stimulation of suppressor lymphocytes contained within the OKT8+ subpopulation.

The effect of angiotensin II on human mononuclear cell reactivity: suppression of PHA-P-induced thymidine incorporation.

Recent evidence suggests that angiotensin II may participate in the regulation of inflammation. We therefore studied the effects of angiotension II on human peripheral blood mononuclear cell reactivity. Peripheral blood mononuclear cell suspensions stimulated with PHA-P revealed decreased thymidine incorporation when simultaneously cultured for 48 or 72 hours with angiotensin II. Experiments were next conducted to determine whether angiotensin II acted directly on lymphocytes. Complete monocyte depletion (greater than 99.5%) resulted in low PHA-P-induced reactivity which was still inhibited by the addition of angiotensin II. Purified (greater than 99.5%) lymphocytes incubated with angiotensin II prior to reconstitution of mixed mononuclear cell suspensions led to reduced PHA-P-stimulated thymidine incorporation. These findings suggest that angiotensin II can effect mononuclear cell uptake possibly through actions on the lymphocyte population.

The effects of indomethacin on in vitro peripheral blood mononuclear cell reactivity in human schistosomiasis.

The peripheral blood mononuclear cell (PBMN) proliferative responses of cells from patients with schistosomiasis were studied in the presence and absence of indomethacin in the culture medium. PBMN cultures were exposed to antigenic extracts of either adult S. mansoni worms (SWAP) or cercariae (CAP), and assayed for the incorporation of tritiated thymidine. More than 70% of the 48 patients studied with SWAP and the 40 patients studied with CAP, were not substantially effected by the addition of indomethacin to the cultures. The remainder (less than 30%) was augmented more than 50% by indomethacin and comprise a group which gave initially low responses to these antigenic preparations. Further analysis indicated that in some schistosomal patients the effect of an adherent suppressor cell population may, in part, be based on a prostaglandin-mediated, indomethacin-sensitive suppressive mechanism. However, the majority of patients, most of whom display adherent suppressor cells, are unaffected by indomethacin. Apparently, other adherent cell suppressor mechanisms are responsible for the regulation observed.