Serum Concentrations Of Proinflammatory Cytokines Research Articles

Flow-cytometric Analysis of Reactive Oxygen Species in Blood Cells: A Potential Tool for Predicting Restenosis - Insights from a Cohort Study.

In-stent restenosis (ISR) is a recurrence of a blockage in a section of the coronary artery that has previously been treated with a stent. Molecular/biochemical pathways underlying ISR are not fully understood, but inflammation and reactive oxygen species (ROS) induced oxidative stress play a significant role in the pathogenesis of restenosis. As blood cells are highly sensitive to oxidative stress and blood is readily accessible compared to other tissues, the current study flow cytometrically investigated intracellular ROS and cytokine profile of blood cells as possible markers of restenosis. Flow cytometry is commonly used for detecting ROS and analyzing oxidative stress but so far, it has not been utilized for prediction of ISR. So, the aim of the study was to explore the potential of flow cytometric assessment of ROS levels in the blood cells as predictor of ISR. The study was carried out in a total of 60 patients who had previously undergone coronary artery stent implantation. They were categorized as Group I - Coronary stent implanted patients without restenosis (n=30) and Group II - Coronary stent implanted patients with restenosis (n=30). Sociodemographics, biochemical and angiographic characteristics were assessed. Intracellular ROS and cytokine estimation in blood cells was done by using flow cytometric analysis. Flow cytometric measurements demonstrated a 1.3-fold increase in ROS levels in red blood cells (RBCs) and 2-fold increase in ROS levels in leucocytes in group II as compared to group I. Mean serum concentrations of pro-inflammatory cytokines: tumor necrosis factor-α (33.54 ± 6.48 vs. 20.10 ± 5.61, p <0.001***), interferon-gamma (21.76 ± 4.46 vs. 20.10 ± 5.61, p <0.001***), interleukin 6 (152.56 ± 30.67 vs. 113.95 ± 23.38, p <0.001***) were found to be higher in restenotic patients as compared to the non-restenotic patients. Correlation analysis showed that intracellular ROS levels of RBCs exhibited a significant positive correlation with late lumen loss in restenotic (r=0.71, p <0.01) as well as non-restenotic patients (r=0.59, p <0.01). Similarly, intracellular ROS levels of WBCs exhibited a significant positive correlation with late lumen loss in restenotic (r=0.72, p <0.01) as well as non-restenotic patients (r=0.61, p <0.01). This study highlights the role of increased levels of intracellular ROS in blood cells in the subsequent development of ISR, which can be detected flow cytometrically. The study suggests that intracellular ROS estimation in blood cells may serve as a potential marker for restenosis and their flow cytometric analysis may facilitate the prediction of ISR.

Dexamethasone administration restored growth in dairy calves exposed to heat stress

Recent evidence indicates that the heat stress loss on the growth performance of calves is associated with the diversion of nutrients to control enteritis and systemic inflammation. In this study, we investigated the impact of heat stress on markers of inflammation, feed use-efficiency, and growth of dairy calves. We hypothesized that dexamethasone, which is known for its immunosuppressive and anti-inflammatory properties, would reduce inflammation and restore the growth of calves exposed to heat stress. Thirty-two Holstein bull calves (body weight (BW) 68.5 ± 1.37 kg; age 3.5 ± 0.5-week-old; mean ± SD) were housed in individual pens in climate-controlled rooms at constant ambient temperature and allowed to adjust to facilities for 5 d before the start of treatments. Calves were randomly assigned to one of 4 treatments (n = 8/treatment) in a 2 × 2 factorial arrangement of environment (ENV, thermoneutral or heat stress) and intervention (INT, saline or dexamethasone) imposed for 5 d as follow: 1) thermoneutral (constant ambient temperature of 20°C 24 h/d) and administration of saline, 2) thermoneutral (constant ambient temperature of 20°C 24 h/d) and administration of dexamethasone, 3) cyclic heat stress (40°C ambient temperature, from 0800 to 1900 h/d) and administration of saline, 4) cyclic heat stress (40°C ambient temperature, from 0800 to 1900 h/d) and administration of dexamethasone. Dexamethasone (0.05 mg/kg BW), or saline (1.2 mL) was administered intramuscularly on d 1 and 3. Upon completion of treatments, calves were euthanized on d 5 to obtain jejunum mucosa samples. Commercial milk replacer, starter grain, and water were offered, and intake was monitored daily. Rectal temperature and respiratory rate were monitored 3 times daily. Blood samples were collected on d 1, 3, and 5 to determine serum pro-inflammatory cytokine concentrations. A section of the jejunum was collected and snap-frozen to determine the concentration of pro-inflammatory markers. Statistical analyses included a mixed model, fixed effects of ENV, INT, consecutive measurements taken over time (d, h, or both), replica, and random effects of calf and error (SAS version 9.4, SAS Institute Inc., Cary, NC). The measurements collected immediately before treatment allocation were included as covariates in the model. An ENV effect showed that heat stress increased rectal temperature (38.72 vs. 39.21°C), respiratory rate (36 vs. 108 breaths/min), and water intake (3.2 vs. 6.6 L/d). The treatments did not affect dry matter intake. An ENV × INT interaction showed that heat stress with saline decreased average daily gain (ADG) by 35% and tended to decrease feed use-efficiency by 36%, but the use of dexamethasone to treat heat stress restored ADG and feed use-efficiency comparable to their basal levels. An ENV × INT interaction revealed that heat stress with saline increased jejunal interleukin (IL)-6 concentration 2-fold, but dexamethasone treatment of heat stress restored jejunal IL-6 concentration to basal levels. The bioenergetic cost of the heat stress-immune pro-inflammatory response ranged between 1.18 and 1.50 Mcal of ME. Overall, the administration of dexamethasone reduced the jejunal concentration of a pro-inflammatory marker and restored the heat stress-associated reduction in growth and feed use-efficiency. The immunomodulation and anti-inflammatory effects of dexamethasone could be part of a homeorhetic change that results in a shift from maintenance functions to support growth on calves exposed to heat stress.

Impact of dietary arginine supplementation on immune responses and growth performance in Newcastle disease virus-infected broiler chicks.

Newcastle disease (ND) poses significant challenges within the poultry industry, leading to increased mortality rates, compromised growth, weakened immunity and elevated levels of inflammation. This study explores the potential of dietary arginine supplementation to ameliorate these adverse effects of ND, leveraging arginine's well-documented benefits in enhancing growth and immune responses. A total of 480 one-day-old male broiler chicks were meticulously categorised into eight groups, encompassing both infected and noninfected cohorts. These chicks received diets with arginine levels at 85%, 100%, 125% and 150% of recommended standards. The study entailed a comprehensive examination of clinical manifestations, growth performance metrics, haemagglutination inhibition (HI) test results, and serum concentrations of proinflammatory cytokines, adrenocorticotropic hormone (ACTH), and cortisol (CORT). The infection significantly curtailed feed consumption (p = 0.0001) and weight gain (p = 0.0001) while concurrently depressing HI titres. Additionally, infected chicks experienced an exacerbated feed conversion ratio (p = 0.0001), escalated mortality rates (p = 0.0001), and elevated serum concentrations of proinflammatory cytokines (p = 0.0001), ACTH (p = 0.0001), and CORT (p = 0.0001). Remarkably, dietary arginine supplementation effectively mitigated the adverse impacts of ND infection on growth, immune responses and proinflammatory cytokine levels. In the context of ND infection, mortality rates and inflammation surge, while growth and immunity are significantly compromised. The strategic inclusion of arginine in the diet emerges as a potent strategy to counteract the deleterious effects of ND. Supplementation with arginine at levels exceeding the conventional dietary recommendations is recommended to alleviate the detrimental consequences of ND effectively.

Therapeutic potential of Pien Tze Huang in colitis-associated colorectal cancer: mechanistic insights from a mouse model

BackgroundPien Tze Huang (PZH), a traditional Chinese medicine formulation, is recognized for its therapeutic effect on colitis and colorectal cancer. However, its protective role and underlying mechanism in colitis-associated colorectal cancer (CAC) remain to be elucidated.MethodsA CAC mouse model was established using AOM/DSS. Twenty mice were randomly divided into four groups (n = 5/group): Control, PZH, AOM/DSS, and AOM/DSS + PZH groups. Mice in the PZH and AOM/DSS + PZH group were orally administered PZH (250 mg/kg/d) from the first day of experiment, while the control and AOM/DSS group received an equivalent volume of distilled water. Parameters such as body weight, disease activity index (DAI), colon weight, colon length, colon histomorphology, intestinal tumor formation, serum concentrations of pro-inflammatory cytokines, proliferation and apoptosis in colon tissue were assessed. RNA sequencing was employed to identify the differentially expressed transcripts (DETs) in colonic tissues and related signaling pathways. Wnt/β-Catenin Pathway-Related genes in colon tissue were detected by QPCR and immunohistochemistry (IHC).ResultsPZH significantly attenuated AOM/DSS-induced weight loss, DAI elevation, colonic weight gain, colon shortening, histological damage, and intestinal tumor formation in mice. PZH also notably decreased serum concentration of IL-6, IL-1β, and TNF-α. Furthermore, PZH inhibited cell proliferation and promote apoptosis in tumor tissues. RNA-seq and KEGG analysis revealed key pathways influenced by PZH, including Wnt/β-catenin signaling pathway. IHC staining confirmed that PZH suppressed the expression of β-catenin, cyclin D1 and c-Myc in colonic tissues.ConclusionsPZH ameliorates AOM/DSS-induced CAC in mice by suppressing the activation of Wnt/β-catenin signaling pathway.

Aberrant Oxygen Concentrations Induce Systemic Inflammation in a Murine Model

Hypoxia is a significant cause of secondary insult in the critically ill trauma or surgical patient. The cause of increased mortality following a brief period of hypoxia is not well understood. The aim of this study is to determine the effect of acute, isolated deviations in oxygen concentration on proinflammatory cytokine release and markers of endothelial stress in a murine model. Mice were randomized to either control, hypoxia, or hyperoxia group. The control group was exposed to room air for 60min, the hyperoxia group was exposed to 70% fraction of inspired oxygen, and the hypoxia group was exposed to 10% fraction of inspired oxygen for 60min. Whole blood collection was completed via cardiac puncture. Serum concentrations of proinflammatory cytokines and endothelial stress markers were analyzed via enzyme-linked immunosorbent assay. Following exposure to hypoxic conditions, there was a significant increase in interleukin (IL)-1α (IL-1 α), IL-1 β, IL-3, IL-4, IL-6, IL-10, tumor necrosis factor α . Following exposure to hyperoxic conditions, there was a significant increase in monocyte chemoattractant protein-1 and regulated upon activation normal T cell expressed and presumably secreted, as well as a significant decrease in IL-12, and IL-17. No clinically significant difference was noted in serum concentration of endothelial stress markers between the treatment groups. Exposure to oxygen extremes induces systemic inflammation as measured by proinflammatory cytokines in a murine model. Hyperoxia also demonstrates the ability to downregulate certain inflammatory pathways while inducing others. No effect on serum concentration of endothelial stress markers is observed following acute, isolated hypoxic or hyperoxic conditions.

LPS-related muscle loss is associated with the alteration of Bacteroidetes abundance, systemic inflammation, and mitochondrial morphology in a weaned piglet model.

We previously demonstrated that lipopolysaccharide (LPS) injection-induced immune stress could impair muscle growth in weaned piglets, but the precise mechanisms behind this remain elusive. Here, we found that chronic immune stress induced by LPS resulted in a significant reduction of 36.86% in the total muscle mass of piglets at 5 d post-treatment compared with the control group. At 1 d, prior to muscle mass loss, multiple alterations were noted in response to LPS treatment. These included a reduction in the abundance of Bacteroidetes, an increase in serum concentrations of pro-inflammatory cytokines, compromised mitochondrial morphology, and an upregulation in the expression of dynamin-related protein 1 (Drp1), a critical protein involved in mitochondrial fission. We highlight a strong negative correlation between Bacteroidetes abundance and the levels of serum pro-inflammatory cytokines, corroborated by in vivo intervention strategies in the musculature of both pig and mouse models. Mechanistically, the effects of Bacteroidetes on inflammation and muscle mass loss may involve the signaling pathway of the tauro-β-muricholic acid-fibroblast growth factor 15. Furthermore, the induction of overexpression of inflammatory cytokines, achieved without LPS treatment through oral administration of recombinant human IL-6 (rhIL-6), led to increased levels of circulating cytokines, subsequently causing a decrease in muscle mass. Notably, pre-treatment with Mdivi-1, an inhibitor of Drp-1, markedly attenuated the LPS-induced elevation in reactive oxygen species levels and rescued the associated decline in muscle mass. Collectively, these data indicate that LPS-induced muscle mass loss was linked to the reduction of Bacteroidetes abundance, increased inflammation, and the disruption of mitochondrial morphology. These insights offer promising avenues for the identification of potential therapeutic targets aimed at mitigating muscle mass loss.

Abstract 6343: AnTenGagerTM, a novel “2+1” T cell engager platform, enables conditional T cell activation with reduced risk of CRS

Abstract Background: CD3+ T cell engaging bispecific antibodies (TCE) redirect T cells to attack targeted tumor cells by simultaneously binding TAA on cancer cells and CD3 complex on a T cell to form a TCR-independent immune synapse. Multiple TCEs have demonstrated promising therapeutic efficacy in the treatment of hematological malignancies. In contrast, suboptimal efficacy in the treatment of solid tumors and the risk of inducing cytokine release syndrome (CRS) continue to be major challenges. Several factors, such as format, affinity, and valency of the antibody, TAA copy number, and antigen size and epitope, can impact the therapeutic efficacy and safety of a TCE. In this study, we developed a novel “2+1” TCE platform, AnTenGagerTM, which enables conditional T cell binding and activation upon TAA-crosslinking, potent anti-tumor activity, and reduced risk of CRS. Methods: Humanized CD3 antibodies were derived from mouse hybridoma, followed by affinity optimization utilizing Computer Aided Drug Design. The Fab-scFv-Fab format was used as the backbone structure of AnTenGagers. The anti-CD3 scFv sequences and the length of peptide linkers between VH/VL, scFv/CH1, and scFv/CH2 were screened and compared in different preclinical assays. LALA mutations (L234A, L235A) were introduced to abolish the Fc receptor binding capability. In a series of in vitro and/or in vivo models, the efficacy, safety and developability of AnTenGagers targeting multiple TAAs, such as Her2, GD2, CDH6, LILRB4, FLT3 and GPRC5D, were evaluated. Results: A library of humanized parental CD3 antibodies with a broad spectrum of affinities were generated, some of which cross react with monkey CD3. AnTenGagers exhibited limited binding to CD3-positive cells before TAA-crosslinking, suggesting a reduced risk of CRS that is caused by systemic CD3+T cell activation. Four CD3 sequences with varying affinities were utilized to construct AnTenGagers with identical TAA-targeting arm. Compared to clinical benchmarks targeting the same TAA, all AnTenGagers demonstrated substantially lower binding capacity to CD3+ cells in the absence of TAA-crosslinking, while inducing increased cytotoxicity against target-positive tumor cells. In addition, they induced significantly less ex-vivo cytokine release by human PBMC than benchmarks. Notably, all AnTenGagers demonstrated enhanced in vivo efficacy compared to the benchmark, exhibiting complete response (CR) in PBMC humanized MM1S mouse CDX model while the serum concentration of proinflammatory cytokines was substantially lower in groups treated with AnTenGagers. AnTenGagers also demonstrated good developability with high expression yield, good thermostability and high stability under different stress conditions. Conclusions: These results suggested that AnTenGagerTM is a promising platform for developing the next generation of TCEs with enhanced efficacy and safety. Citation Format: Bing Hou, Hui Yuwen, Tengteng Li, Yijing Ren, Ao Sun, Zaoshun Hu, Jay Mei, Bo Shan. AnTenGagerTM, a novel “2+1” T cell engager platform, enables conditional T cell activation with reduced risk of CRS [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6343.

Relationship between biomarkers of autophagy and inflammation in acute period of ischemic stroke

Postischemic neuroinflammation is a critical pathophysiological process within the entire pattern of cerebral ischemia, spanning early injury and tissue repair. According to recent experimental data, autophagy is involved in the regulation of neuroinflammation, influencing the outcome of the acute period of ischemic stroke (IS).Objective. To evaluate the relationship between autophagy biomarkers and inflammation indicators in the dynamics of the acute period of atherothrombotic IS.Materials and methods. 112 patients in the acute period of newly developed atherothrombotic IS and 56 donors (control group) were examined. Patients underwent dynamic clinical and neurological examination on the 1st, 7th and 14th days from the onset of the disease (magnetic resonance imaging, testing using the NIHSS scale, modified Rankin scale). At the same time intervals, blood was drawn for testing. The number of active autophagosomes in peripheral blood was assessed by flow cytometry using a specific Cyto-ID dye. The serum concentrations of proinflammatory cytokines IL‑1β, IL‑8, IL‑18 (interleukins‑1β, -8, -18), TNFα (tumor necrosis factor-α), autophagy biomarkers Beclin‑1, LC 3 and p62 were determined by enzyme-linked immunosorbent assay analysis. C-reactive protein was assessed by a highly sensitive immunoturbidimetric method.Results. A statistically significant increase in the studied parameters was revealed compared to the control group. The maximum increase in inflammation biomarkers was observed on the 1st day, and the maximum increase in key indicators of autophagy (LC 3, Beclin‑1, Cyto-ID) – on the 7th day after the development of ischemia. A direct relationship was established between the level of autophagy and the concentration of inflammatory biomarkers (CRP, IL‑1β, IL‑18, TNF-α) on the 1st and 7th days of acute IS.Conclusions. The identified correlations indicate the participation of activated autophagy in the regulation of post-ischemic neuroinflammation and its involvement in ischemic brain damage in the early stages of the acute period of IS (days 1–7). The results obtained confirm the literature data on the influence of autophagy on the outcome of the acute period of the disease.

Pioneering PGC-1α-boosted secretome: a novel approach to combating liver fibrosis.

Liver fibrosis is a critical health issue with limited treatment options. This study investigates the potential of PGC-Sec, a secretome derived from peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α)-overexpressing adipose-derived stem cells (ASCs), as a novel therapeutic strategy for liver fibrosis. Upon achieving a cellular confluence of 70%-80%, ASCs were transfected with pcDNA-PGC-1α. PGC-Sec, obtained through concentration of conditioned media using ultrafiltration units with a 3-kDa cutoff, was assessed through in vitro assays and in vitro mouse models. In vitro, PGC-Sec significantly reduced LX2 human hepatic stellate cell proliferation and mitigated mitochondrial oxidative stress compared to the control-secretome. In an in vivo mouse model, PGC-Sec treatment led to notable reductions in hepatic enzyme activity, serum proinflammatory cytokine concentrations, and fibrosis-related marker expression. Histological analysis demonstrated improved liver histology and reduced fibrosis severity in PGC-Sec-treated mice. Immunohistochemical staining confirmed enhanced expression of PGC-1α, optic atrophy 1 (a mitochondrial function marker), and peroxisome proliferator-activated receptor alpha (an antifibrogenic marker) in the PGC-Sec-treated group, along with reduced collagen type 1A expression (a profibrogenic marker). These findings highlight the therapeutic potential of PGC-Sec in combating liver fibrosis by enhancing mitochondrial biogenesis and function, and promoting antifibrotic processes. PGC-Sec holds promise as a novel treatment strategy for liver fibrosis.

Glycoprotein 5-Derived Peptides Induce a Protective T-Cell Response in Swine against the Porcine Reproductive and Respiratory Syndrome Virus.

We analyzed the T-cell responses induced by lineal epitopes of glycoprotein 5 (GP5) from PRRSV to explore the role of this protein in the immunological protection mediated by T-cells. The GP5 peptides were conjugated with a carrier protein for primary immunization and booster doses. Twenty-one-day-old pigs were allocated into four groups (seven pigs per group): control (PBS), vehicle (carrier), PTC1, and PTC2. Cytokine levels were measured at 2 days post-immunization (DPI) from serum samples. Cytotoxic T-lymphocytes (CTLs, CD8+) from peripheral blood were quantified via flow cytometry at 42 DPI. The cytotoxicity was evaluated by co-culturing primed lymphocytes with PRRSV derived from an infectious clone. The PTC2 peptide increased the serum concentrations of pro-inflammatory cytokines (i.e., TNF-α, IL-1β, IL-8) and cytokines that activate the adaptive cellular immunity associated with T-lymphocytes (i.e., IL-4, IL-6, IL-10, and IL-12). The concentration of CTLs (CD8+) was significantly higher in groups immunized with the peptides, which suggests a proliferative response in this cell population. Primed CTLs from immunized pigs showed cytolytic activity in PRRSV-infected cells in vitro. PTC1 and PTC2 peptides induced a protective T-cell-mediated response in pigs immunized against PRRSV, due to the presence of T epitopes in their sequences.

Effects of mycobacterium cell wall fraction on embryo development following in vitro embryo production and pregnancy rates following embryo transfer in virgin dairy heifers

An experiment was conducted to determine whether administration of mycobacterium cell wall fraction (MCWF; Amplimune, NovaVive) could enhance embryo developmental competence following in vitro embryo production (IVP) and pregnancy establishment after embryo transfer (ET). Nulliparous, Holstein heifers (n=40; age 8-15 months) were submitted to two rounds of ovum pick-up (OPU) and IVP in a crossover design. Thirty-six h after follicle wave synchronization, treatments (saline or MCWF, 5mL, im) were administered in conjunction with a single dose of follicle stimulating hormone (175 IU) and OPU was performed 48-52h later. Recovered cumulus-oocyte complexes were used for IVP to assess embryo development. For ET, nulliparous, Holstein heifers (n=225; age 12-18 months) were used as recipients. At 12-24h after detection of spontaneous estrus, recipients were randomly treated with either saline or MCWF (5mL, im). The effect of MCWF on pregnancy per ET (P/ET) was assessed in a 2×2 factorial design with recipients treated with or without MCWF receiving a fresh IVP embryo from a donor treated with or without MCWF at day 7 or 8 after detected estrus. Blood samples were collected from a subset of donors (n=8) and recipients (n=26 to 33 per treatment) prior to treatment and at 6 and 24h post-treatment to determine serum concentration of interleukin (IL)-1β, IL-6, tumor necrosis factor-α, and interferon-γ. Blood samples were also collected from a group of recipients (n=31 to 39 per treatment) to assess serum concentration of progesterone at days 4, 7, and 16 post-treatment. Pregnancy status was determined at days 40 and 100 of gestation. Donor treatment with MCWF tended (P<0.07) to increase the proportion of oocytes that developed into transferable embryos, but there was no effect of MCWF on other parameters of embryo development. The P/ET at days 40 and 100 of gestation and pregnancy loss were not affected by donor treatment or recipient treatment with MCWF and there was no interaction. Serum concentration of proinflammatory cytokines among donors and recipients and serum concentration of progesterone among recipients were not increased by treatment with MCWF. Results of the present study indicate that treatment of donors with MCWF has minimal impact on subsequent embryo development following IVP. Moreover, regardless of whether donors or recipients were treated with MCWF, there was no effect on P/ET following transfer of IVP embryos.

Pro-inflammatory cytokines and reproductive hormone responses in bucks post-challenge with Mannheimia haemolytica A2 and its outer membrane protein.

The lipopolysaccharide (LPS) endotoxin and outer membrane protein (OMP) are among the virulence factors of Gram-negative bacteria responsible for inducing pathogenicity in the infected host. OMP and LPS occur on the outer membrane of M. haemolytica A2, the primary aetiological agent of pneumonic mannheimiosis in small ruminants. While the LPS is known to mediate Gram-negative bacterial infection by activating downstream inflammatory pathways, the potential role of OMP during inflammatory responses remained unclear. Hence, this study determined the effect of the OMP of M. haemolytica A2 on the serum concentration of pro-inflammatory cytokines and the male reproductive hormones (testosterone and Luteinizing Hormone). We randomly assigned twelve bucks to three groups (n = 4 bucks each): Group 1 was challenged with 2mL PBS buffer (pH 7.0) intranasally; Group 2 received 2mL of 1.2 X 109CFU/mL whole M. haemolytica A2 intranasally; and Group 3 received 2mL of OMP extract obtained from 1.2 X 109CFU/mL M. haemolytica A2 intramuscularly. Serum samples collected at pre-determined intervals were used for the quantitative determination of the pro-inflammatory cytokines (IL-1β, IL-6, and TNFα) and reproductive hormones (testosterone and LH) using commercial sandwich enzyme-linked immunosorbent assay (ELISA). The serum concentration of IL1β was initially increased within the first-hour post-challenge in Groups 2 and 3, followed by a significant decrease in concentration at 21d and 35d (p < 0.05) in Group 3. Only mild fluctuations in IL-6 occurred in group 2, as opposed to the 1.7-fold rapid increase in TNFα within 2h post-challenge before decreasing at 6h. An increase in pro-inflammatory cytokines was accompanied by an acute febrile response of 39.5 ± 0.38°C (p < 0.05) at 2h and 40.1 ± 0.29°C (p < 0.05) at 4h in Group 2 and Group 3, respectively. Serum testosterone decreased significantly (p < 0.05) in both treatment groups but remained significantly (p > 0.05) lower than in Group 1 throughout the study. There was a moderate negative association between testosterone and IL1β (r = -0.473; p > 0.05) or TNFα (r = -0.527; p < 0.05) in Group 2. Serum LH also showed moderate negative associations with TNFα in Group 2 (r = -0.63; p < 0.05) and Group 3 (r = -0.54; p > 0.05). The results of this study demonstrated that M. haemolytica A2 and its OMP produced marked alterations in serum levels of pro-inflammatory cytokines and male reproductive hormones. The negative correlations between serum testosterone and inflammatory cytokines would suggest the potential role of OMP in causing male infertility by mediating innate inflammatory responses to suppress testosterone production in bucks.

Characterization of the inflammatory response to COVID-19 illness in pregnancy

Pregnant patients face greater morbidity and mortality from COVID-19 related illness than their non-pregnant peers. Previous research in non-pregnant patients established that poor clinical outcomes in SARS-CoV-2 positive patients admitted to the ICU were correlated with a significant increase in the proinflammatory markers interleukin (IL)-1β, IL-6, IL-8, and IL-10. Importantly, high levels of these inflammatory markers have also been associated with adverse pregnancy outcomes, including spontaneous preterm birth, preeclampsia, and severe respiratory disease. This was a retrospective cohort study that compared the serum inflammatory cytokine profiles of pregnant patients with acute/post-acute SARS-CoV-2 infection to those with previous exposure. All subjects in both cohorts tested positive for SARS-CoV-2 antibodies; however, those in the acute/post-acute infection cohort had a documented positive SARS-CoV-2 reverse transcription polymerase chain reaction (RT-PCR) result within 30days of serum sample collection. Serum samples were obtained during prenatal venipuncture from 13 to 39weeks' gestation and the cohorts were matched by gestational age. The inflammatory cytokines interferon (IFN)-γ, IL-10, IL-1β, IL-4, IL-6, IL-8, and tumor necrosis factor (TNF)-α were assayed from maternal serum using a standard ELISA assay and median cytokine concentrations were compared using the Mann-Whitney test. We enrolled 50 non-Hispanic Black patients with confirmed COVID-19 infection who received prenatal care at Grady Memorial Hospital in Atlanta, Georgia. Those with acute/post-acute infection (n=22) had significantly higher concentrations of SARS-CoV-2 antibody, IL-10, IL-1β, and IL-8, while patients with previous exposure (n=28) had significantly higher concentrations of IL-4. There were no significant inter-group differences in medical comorbidities. Pregnant patients with acute/post-acute SARS-CoV-2 infection had significantly higher serum concentrations of pro-inflammatory cytokines as compared to those with previous exposure, suggesting that, like in the non-pregnant population, SARS-CoV-2 infection alters the levels of circulating proinflammatory markers during pregnancy. The increased levels of cytokines may contribute to the adverse obstetric outcomes observed with COVID-19 illness.

A Pilot Study on Burnout in Medical Students (BuMS) over an Academic Year

Background: Physician burnout is increasingly recognized as a problem in physician well-being and may negatively affect patient care outcomes. Burnout can begin at any point of training or practice, potentially as early as the first year of medical school. Thus, there is a need to characterize possible burnout in medical students as the first step to optimizing strategies for mitigation. Traditionally, burnout has been studied using survey-based variables; however, identifying novel physiological and molecular biomarkers could allow for the expansion of screening and intervention strategies. Methods: In this pilot prospective cohort study, we followed a group of preclinical 1st and 2nd year medical students (n = 9) at the University of Florida over one academic year of medical school. We collected survey responses (Maslach Burnout Inventory [MBI], Patient Health Questionnaire-9 [PHQ-9], and Perceived Stress Scale [PSS]) and measured a panel of candidate physiological biomarkers of burnout (Inflammatory Cytokine Panel, Heart Rate Variability [HRV], and Leukocyte Telomere Length). Results: In the study participants, MBI composite scores and PHQ-9 scores showed a statistically significant increase over the course of an academic year, indicating higher levels of medical student burnout. Additionally, respondents reported a statistically significant decrease in time devoted to exercise, and we measured a significant increase in body mass index (BMI) during the academic year. PSS scores showed an upward trend which was not statistically significant. Likewise, average leukocyte telomere length trended downward, but the change was not statistically significant. There were no measured changes in the serum concentration of pro-inflammatory cytokines, and time-domain heart rate variability metrics did not differ significantly between timepoints. Conclusions: This pilot study supports the notion that burnout can begin early in medical school and is detectable via survey instruments in first-year and second-year medical students even with a small sample size. Additionally, leukocyte telomere length could potentially be a useful biomarker of burnout with supporting data, but we did not observe any statistically significant changes in inflammatory cytokines or heart rate variability. Further investigation into these potential biomarkers with larger cohort sizes is required to fully characterize their clinical utility.

Assessment of gastrointestinal health in racing Alaskan sled dogs using capsule endoscopy and inflammatory cytokines.

Exercise-induced gastrointestinal syndrome occurs in dogs and people and might compromise athlete performance by increasing intestinal permeability and causing gastrointestinal erosions. Racing sled dogs often receive acid suppressant prophylaxis which decreases the incidence of gastric erosions induced by exercise. The objectives were to quantify intestinal injury by measuring serum pro-inflammatory cytokine concentrations before and after exercise and to evaluate gastrointestinal mucosa using video capsule endoscopy after exercise. Prospective study of 12 racing Alaskan sled dogs receiving approximately 1 mg/kg omeprazole once daily from the day before the race until race completion. Blood was drawn before and 8 to 10 hours after an endurance race for the quantification of cytokines. Gastrointestinal tract mucosa was assessed with video capsule endoscopy immediately post-race. Eight of nine dogs (89%; 95% confidence interval 52 to 100%) had gastric erosions; all dogs (100%, 95% confidence interval 63 to 100%) had small intestinal erosions. Most of the dogs (seven of nine) had straw or foreign material present. Cytokine levels were not different from before to after the race. Video capsule endoscopy identified gastrointestinal tract mucosal erosions after exercise in all dogs receiving once-daily omeprazole treatment, though other causes for the lesions besides exercise are possible.

CAR +and CAR −T cells differentiate into an NK-like subset that is associated with increased inflammatory cytokines following infusion

Abstract Chimeric antigen receptor (CAR) T cells have demonstrable efficacy in treating B-cell malignancies. Factors such as product composition, lymphodepletion and immune reconstitution are known to influence CAR +T cell survival and persistence, however the determinants of CAR +T cells’ fate and differentiation and how the patients’ reconstituting immune system influence therapy outcomes remain poorly understood. We applied single cell multi-omics analysis of RNA, protein and clonal profiles of CAR +T and PBMC in patients receiving donor-derived products from a piggybac CAR19 clinical trial (N=8 patients). We used these data to reconstruct a differentiation trajectory, which explained the observed phenotype and fate of CAR +and CAR −T cells within the first 30 days of post-infusion. Following initial lympho-depletion, endogenous CAR −CD8 +, γδ T cells and CAR+ T cells clonally expanded, and differentiated across heterogenous phenotypes. They transitioned from a dominant resting or proliferating state into precursor of exhausted T cells, and finally into a terminal NK-like phenotype. The subset of terminal NK-like CAR +T cells was associated with increased serum concentrations of proinflammatory cytokines including IL-12 and IL-18. Validation on a published cohort (61 patients) showed that this subset was associated with clinical outcome at 6 months. Finally, we analysed in detail one patient that developed CAR T cell induced lymphoma and discussed the impact of early phase therapy in determining this unexpected outcome. These results demonstrate that CAR- and CAR+ T cells share a differentiation profile which may be determined by non-CAR derived signals and which may influence patients’ immune-recovery and outcome. National Health And Medical Research Funding (Australia) (APP1121643, APP1128416) BD Bioscience

Efficiency of conventional and nanoparticle oxytetracycline in treatment of clinical endometritis in postpartum dairy cows

The incidence of clinical endometritis in dairy cows postpartum is one of the important reasons for financial losses in the dairy industry. The costs of treatment, milk losses, infertility, repeated breeding, and high annual culling rate of dairy cows present immediate losses in case of treatment failure. The commonly used therapeutic methods for clinical endometritis have not been successful nor have given definitive solutions to overcome the complications of the disease in dairy cows. Therefore, it was necessary to propose an innovative treatment program to overcome the reasons for the failure and lack of effectiveness of the treatment of clinical endometritis. This was tackled in the current study; oxytetracycline with different concentrations, oxytetracycline 5% (OTCC5%), oxytetracycline 20% (OTCC20%), and oxytetracycline 20% nanoparticles (OTC-NPs) were used for the treatment of clinical endometritis. Diagnosis of clinical endometritis was based on the assessment of high serum concentration of pro-inflammatory cytokines, acute phase protein, increased endometrium thickness, and intrauterine discharges with different degrees of echogenicity monitored by ultrasonography. Application of OTC-NPs revealed a decrease in serum concentration of pro-inflammatory cytokines (IL-1, IL-6, and TNF-α) and acute phase proteins compared to OTCC20% and OTCC5% groups. The improvement achieved by OTC-NPs may be attributed to the reduction of OTC particles into nano size which facilitates its tissue bioavailability, dispersion, penetration power to deeper tissues, and its more broad-spectrum activities. These activities were clearly apparent after the evacuation of uterine contents using a single dose of PGF2α. The OTC-NPs revealed a reduction in serum concentration of cytokines compared to OTCC20% and OTCC5% groups arranged as follows: 10.11, 25.45, 35.56 for IL-1; 99, 300, 319 for IL-6; 1.01, 4.40, 8.06 for CRP; and 46, 183, 266 for TNF-α. Furthermore, an increase in serum concentration of albumin (3.34) was obtained by OTC-NPs compared to OTCC5% (1.70). This improvement can be taken as evidence of liver resumption functions and inflammatory reactions. On the other side, globulin concentration recorded an increase like albumin and total proteins in OTC-NPs compared to others. A reduction in the endometrium thickness in OTC-NPs with the disappearance of intrauterine discharges was monitored by ultrasonography. This confirmed the subsiding of clinical endometritis in OTC-NPs group. Moreover, a significant improvement in conception and pregnancy rate in OTC-NPs compared to other groups were observed.

Dissolving microneedle patches-mediated percutaneous delivery of tetramethylpyrazine for rheumatoid arthritis treatment

Recently, transdermal treatment of rheumatoid arthritis (RA) has received increasing attention due to the advantages of improving patient compliance and avoiding gastrointestinal side effects. However, the stratum corneum (SC) barrier limits the transdermal delivery of most substances. Therefore, we constructed tetramethylpyrazine-loaded dissolving microneedle patches (TMP-DMNPs) and investigated its anti-rheumatoid arthritis effect. The cone-shaped dissolving microneedle patch had complete, neatly arranged needles and great mechanical strength. It could effectively penetrate the stratum corneum when applied to the skin. In vitro transdermal experiment showed that DMNPs could significantly promote the transdermal penetration of TMP compared with TMP-cream. The needles were completely dissolved within 18min and the applied skin recovered completely within 3h. The excipients and blank DMNP had good safety and biocompatibility to human rheumatoid arthritis fibroblast synovial cells. To compare the therapeutic effects, the animal model was established. The experiments of paw swelling, histopathology and X-ray examination showed that dissolving microneedles significantly alleviated paw condition, reduced the serum concentrations of proinflammatory cytokines, and inhibited synovial tissue damage in AIA rats. These results indicate that the DMNPs we prepared can deliver TMP safely, effectively and conveniently, providing a basis for the percutaneous treatment of RA.

The effects of omega 3 fatty acids on the serum concentrations of pro inflammatory cytokines anddepression status in patients with bipolar disorder: A randomized double-blind controlled clinical trial.

The inflammation accelerates the progression of bipolar disorder. Supplementation of anti-inflammatory supplements in adjuvant with medications may alleviate disorder signs. This study aimed to investigate the effects of omega-3 fatty acid supplementation on the serum concentrations of pro-inflammatory cytokines and depression status in patients with bipolar disorder. This randomized clinical trial study was conducted in Zahedan city in 2021. Patients with bipolar disorder (n = 60) were grouped into two groups: omega-3 fatty acid supplement group (n = 30, 15 men and 15 women) and placebo one using a permuted block stratified randomization. The patients in the omega-3 group received 2 g of omega-3 fatty acids daily for 2 months while patients in the placebo group received 2 g soft gels daily in the same form. Depression score and the serum concentrations of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) and high-sensitivity C-reactive protein (hs-CRP) were assessed before and after the study. Depression score and the serum concentrations of TNF-α, IL-6, and hs-CRP were decreased after intervention in the omega-3 fatty acid group also compared with the placebo group (P < 0.001). The results also show a positive correlation between the serum concentrations of TNF-α, IL-6, and hs-CRP with depression scores (P < 0.001). Prescription of omega-3 fatty acids can decrease inflammatory parameters and help to decrease depression in patients with bipolar disorder. This supplement can be used along with medications for decreasing the inflammatory markers in these patients.

Оптимізація лікування гострого тонзилофарингіту в дитячому віці

Acute tonsillopharyngitis (ATP) in children is an actual and not fully solved problem, which has interdisciplinary importance and requires optimization of diagnosis and treatment to improve the prognosis of the disease and the quality of life of children. Purpose - to optimize the tactics of treatment of children with ATP by supplementing the complex of therapy with drugs of anti-inflammatory and immunomodulatory action with the evaluation of clinical and immunological markers of the effectiveness of the proposed treatment. Materials and methods. We monitored 51 hospitalized children with a clinical diagnosis of ATP. All children underwent a complex clinical and paraclinical examination with the determination of C-reactive protein (CRP) in the blood serum upon admission to the hospital, and on the 7th day of treatment, the serum concentration of interleukin (IL) -4, -6, -8, tumor necrosis factor α (TNFα), and interferon-γ (γ-IFN) as markers of therapy effectiveness. The patients were randomly divided into 2 clinical groups: the Group 1 was formed by 28 patients (average age - 8.0±0.9 years, the share of boys - 53.5±6.1%) with ATP, who received standard treatment and standardized extract of Pelargonium sidoides EPs® 7630 (Umckalor) and the complex homeopathic drug Tonsilotren in age-related doses according to the instructions. The remaining 23 children formed the clinical Group 2 (average age - 8.7±1.0 years (p&gt;0.05), the share of boys - 56.5±8.3% (p&gt;0.05), and received the same standard treatment for ATP and served as a comparison group. The groups were matched for the main clinical characteristics. Results. We obtained significantly better results of treatment of children with ATP when using the combination of the drugs Umckalor and Tonsilotren in complex therapy, which are proven by the presence of statistically reliable chances of a milder clinical course of the disease on the 3rd day of treatment (assessment of the severity of ATP - &lt;26 points): attribute risk - 59.8%, relative risk - 2.8 (95% CI: 1.1-7.2), odds ratio - 25.6 (9.0-72.6). The effectiveness of the proposed therapy was confirmed on the 7th day of treatment by the probably lower content of pro-inflammatory markers IL-8 (15.2±5.4 pg/ml vs 37.6±7.6 pg/ml) and γ-IFN (21.8±2.3 pg/ml vs. 30.0±1.2 pg/ml) compared to standard treatment, as well as twice lower TNF-α concentration (2.1±0.5 pg/ml vs. 4.5±2.5 pg/ml). The use of the combination of the drugs Umckalor and Tonsilotren was accompanied by a statistically significant reduction in the risk of maintaining a high serum concentration of pro-inflammatory cytokines (CRP - &gt;10.0 mg/l, γ-IFN and IL-8&gt;10.0 pg/ml) in relation to the standard treatment: absolute risk reduction - 20.8%, relative risk reduction - 32.3% with the minimum required number of patients to treat - 3.1. Conclusions. In the complex treatment of children with ATP, the use of a combination of the standardized extract of Pelargonium sidoides EPs® 7630 (Umckalor) and the complex homeopathic drug Tonsilotren demonstrated better effectiveness compared to the same standard treatment. The research was carried out in accordance with the principles of the Helsinki Declaration. The study protocol was approved by the Local Ethics Committee of the participating institution. The informed consent of the patient was obtained for conducting the studies. No conflict of interests was declared by the authors.