Immunoregulatory Effects of the Active Form of Vitamin D (Calcitriol), Individually and in Combination with Curcumin, on Peripheral Blood Mononuclear Cells (PBMCs) of Multiple Sclerosis (MS) Patients.

OBJECTIVES: Multiple sclerosis (MS) is a chronic autoimmune inflammatory disease affecting the central nervous system. Immune cell subsets, notably T helper (Th) 17 and Th1, exert important roles in MS pathogenesis. Whereas, Treg cells modulate the disease process. Calcitriol, the active form of vitamin D, and curcumin, a bioactive compound derived from turmeric, play immunomodulatory effects relevant to autoimmune disorders, including MS. The objective of this study is to investigate the effects of calcitriol and Curcumin on Peripheral blood mononuclear cells (PBMCs) of individuals with MS. METHODS: PBMCs from twenty MS patients were isolated, cultured, and exposed to 0.004 µg/mL of calcitriol and 10 µg/mL of curcumin. The cells underwent treatment with singular or combined doses of these components to assess potential cumulative or synergistic immunomod-ulatory effects. Following treatment, the expression levels of genes and the cellular population of Treg, Th1 and Th17 were evaluated using Real-time PCR and flow cytometry. RESULTS: Treatment with curcumin and calcitriol led to a significant reduction in the expression levels of inflammatory cytokines and transcription factors related to Th1 and Th17 cells, includ-ing IFN-γ, T-bet, IL-17, and RORC. Furthermore, the frequency of these cells decreased follow-ing treatment. Additionally, curcumin and calcitriol treatment resulted in a significant upregu-lation of the FOXP3 gene expression and an increase in the frequency of Treg cells. CONCLUSION: This study demonstrates that curcumin and calcitriol can effectively modulate the inflammatory processes intrinsic to MS by mitigating the expression of inflammatory cytokines by Th1 and Th17 cells while concurrently enhancing the regulatory role of Treg cells. Moreover, the combined treatment of curcumin and calcitriol did not yield superior outcomes compared to single-dosing strategies.

The Potential Hepatoprotective Effect of Vaccinium arctostaphylos L. Fruit Extract in Diabetic Rat.

OBJECTIVE: Vaccinium arctostaphylos has traditionally been employed in Iranian folk medicine to treat diabetes. However, the precise molecular mechanisms underlying its antidiabetic properties remain incompletely understood. The current experiment intended to explore the modulatory effects of V. arctostaphylos fruit ethanolic extract (VAE) on biochemical and molecular events in the livers of diabetic rats. MATERIALS AND METHODS: In this experimental study, male Wistar rats were randomly assigned to four groups: normal control, normal rats with VAE treatment, diabetic control, and diabetic rats with VAE treatment. Following 42 days of treatment, the impact of VAE on diabetes-induced rats was assessed by measuring various serum biochemical parameters, including insulin, free fatty acids (FFA), tumor necrosis factor-α (TNF-α), reactive oxygen species (ROS), and adiponectin levels. The activities of hepatic carbohydrate metabolic enzymes and glycogen content were determined. Additionally, expression levels of selected genes implicated in carbohydrate/lipid metabolism and miR-27b expression were evaluated. H and E-stained liver sections were prepared for light microscopy examination. RESULTS: Treatment with VAE elevated levels of insulin and adiponectin that reduced levels of FFA, ROS, and TNF-α in the serum of diabetic rats. VAE-treated rats exhibited increased activities of hepatic glucokinase (GK), glucose-6-phosphate dehydrogenase (G6PD), and glycogen concentrations, in conjunction with decreased activities of glucose-6-phosphatase (G6Pase) and fructose-1,6-bisphosphatase (FBPase). Furthermore, VAE significantly upregulated the transcription levels of hepatic insulin receptor substrate 1 (Irs1) and glucose transporter 2 (Glut2), while considerably downregulated the expression of peroxisome proliferator-activated receptor gamma (Pparg) and sterol regulatory element-binding protein 1c (Srebp1c). VAE remarkably enhanced the expression of miR27-b in the hepatic tissues of diabetic rats. Abnormal histological signs were dramatically normalized in diabetic rats receiving VAE compared to those in the diabetic control group. CONCLUSION: Our findings underscore the hypoglycemic and hypolipidemic activities of V. arctostaphylos and assist in better comprehension of its antidiabetic properties.

Immunosuppressive Effects of Two Probiotics, Lactobacillus paracasei DSM 13434 and Lactobacillus plantarum DSM 15312, on CD4+ T Cells of Multiple Sclerosis Patients.

T cells play an important role in the development and progression of multiple sclerosis (MS), an autoimmune disease of the central nervous system. In the present study, the immunomodulatory impacts of two Lactobacillus strains, L paracasei DSM 13434 and L plantarum DSM 15312, on the frequency and cytokine production of CD4+ T cells in MS patients were explored. Thirty MS patients were enrolled in this study. The CD4+ T cells were isolated, cultured, and exposed to the media containing cell-free supernatants of L plantarum (group1), L paracasei (group 2), the mixture group of cell-free supernatants of both probiotics (group 3), and vehicle (control) group (group 4). The frequencies of T helper (Th) 1, Th17, Th2, and T regulatory type 1 (Tr1) cells and mean fluorescent intensity (MFI) of the associated cytokines were assessed using flow cytometry. The levels of interleukin 17 (IL-17), transforming growth factor ß (TGF-ß), and interferon-gamma (IFN-γ) cytokines in supernatants of all groups were measured by enzyme-linked immunosorbent assay. The percentage of Th1 cells and the MFI of IFN-γ in Th1 cells (CD4+ IFN-γ+) in all three probiotic treatment groups were significantly decreased compared to the control group. However, no significant changes were observed in the proportion and MFI of Th2, Th17, and Tr1 cells. A significant decrease was observed in IL-17 secretion in the supernatant of cultured CD4+ T cells in all three treatment groups in comparison with control. The levels of TGF-ß and IFN-γ were not significantly different among any of the study groups.  Collectively, cell-free supernatants of the lactobacilli showed an in vitro anti-inflammatory effect. However, further studies are needed to prove the real effects of probiotics on MS.

New insights into extracellular and intracellular redox status in COVID-19 patients.

BACKGROUND: The imbalance of redox homeostasis induces hyper-inflammation in viral infections. In this study, we explored the redox system signature in response to SARS-COV-2 infection and examined the status of these extracellular and intracellular signatures in COVID-19 patients. METHOD: The multi-level network was constructed using multi-level data of oxidative stress-related biological processes, protein-protein interactions, transcription factors, and co-expression coefficients obtained from GSE164805, which included gene expression profiles of peripheral blood mononuclear cells (PBMCs) from COVID-19 patients and healthy controls. Top genes were designated based on the degree and closeness centralities. The expression of high-ranked genes was evaluated in PBMCs and nasopharyngeal (NP) samples of 30 COVID-19 patients and 30 healthy controls. The intracellular levels of GSH and ROS/O2• - and extracellular oxidative stress markers were assayed in PBMCs and plasma samples by flow cytometry and ELISA. ELISA results were applied to construct a classification model using logistic regression to differentiate COVID-19 patients from healthy controls. RESULTS: CAT, NFE2L2, SOD1, SOD2 and CYBB were 5 top genes in the network analysis. The expression of these genes and intracellular levels of ROS/O2• - were increased in PBMCs of COVID-19 patients while the GSH level decreased. The expression of high-ranked genes was lower in NP samples of COVID-19 patients compared to control group. The activity of extracellular enzymes CAT and SOD, and the total oxidant status (TOS) level were increased in plasma samples of COVID-19 patients. Also, the 2-marker panel of CAT and TOS and 3-marker panel showed the best performance. CONCLUSION: SARS-COV-2 disrupts the redox equilibrium in immune cells and the upper respiratory tract, leading to exacerbated inflammation and increased replication and entrance of SARS-COV-2 into host cells. Furthermore, utilizing markers of oxidative stress as a complementary validation to discriminate COVID-19 from healthy controls, seems promising.

Alteration of the Inflammatory and Anti-Inflammatory Cytokine Profiles of Peripheral Blood Mononuclear Cell in Crohn's Disease Patients after Following up.

Background: Crohn's disease (CD) has a chronic course, which its recurrence varies widely among different patients. In this study we prospectively analyzed blood samples of 19 CD patients. Alteration in transcription of inflammatory and anti-inflammatory cytokines was analyzed compared with household members after three month follow up. Methods: CD patients were diagnosed based on clinical symptoms, endoscopic and histopathologic characteristics. Nineteen CD patients and their households were evaluated from Jun 2019 to Feb 2021 at Tehran university hospitals. CD activity score, biological, clinical and demographic data of the patients were recorded at two time point intervals. Bacteriological tests were done using aerobic and anaerobic blood cultures. To investigate transcriptional alterations, peripheral blood mononuclear cells (PBMCs) were isolated using Ficol centrifugation method and relative quantitative real-time PCR was done to determine the expression level of IFN-γ, TNF-α, IL10, and FOXP3 cytokines. Results: Our results showed a correlation between fecal calprotectin level (709.8 ± 554.6), C-reactive protein concentration (18.1 ± 15.9), and erythrocyte sedimentation rate (30.4 ± 17.9) with disease activity (Flare/remission). IL10 and Foxp3 anti-inflammatory gene's expression were significantly (P = 0.003 for IL10 and P = 0.008 Foxp3) higher during the flare and remission in patients with active disease respectively. Bacteriological examination showed infection with Streptococcus spp. and Clostridium spp. in two CD patients during flares, which was correlated with upregulation and down-regulation of IL10, TNF-α, IFN-γ and FOXP3 proteins, respectively. Conclusion: Occurrence of bacteremia, and higher amount of CAP, CRP and ESR are correlated with higher level of transcription for inflammatory cytokines, which could effectively reflect the disease activity. Raise in FoxP3 transcription proposed change in Treg sub-population in PBMC or its activity during the CD remission phase.

Mesenchymal stem cells and their conditioned medium as potential therapeutic strategies in managing comorbid anxiety in rat sepsis induced by cecal ligation and puncture.

Objectives: Sepsis-associated encephalopathy (SAE) is a common brain dysfunction following sepsis. Due to the beneficial effects of mesenchymal stem cells (MSCs) therapy on anxiety, an extreme and early manifestation of SAE, we hypothesized that MSCs-derived conditioned medium (CM) may be able to attenuate anxiety in cecal ligation and puncture (CLP)-induced sepsis. Materials and Methods: Rats were assigned into 4 groups: sham, CLP, MSC, and CM. All animals, except in the sham group, underwent the CLP procedure to induce sepsis. Two hours after sepsis induction, the rats in MSC and CM groups, received 1×106 MSCs and CM derived from the same number of cells, respectively. 48 hr after the treatments, anxiety-related behaviors were assessed, and brain and right hippocampal tissues were collected. Results: MSCs and CM enhanced the percentages of open arm entries and time spent in the open arms of the elevated plus-maze and the time spent in the light side of the light-dark box. MSCs and CM decreased the Evans blue content and decreased the IL-6 and TNF-α levels in the brain tissue samples. Reductions in the expression of 5-HT2A receptors and phosphorylation of ERK1/2 and an increase in the expression of 5-HT1A receptors in the hippocampal tissue samples were observed in the MSC and CM groups. Conclusion: MSCs and MSCs-derived CM attenuated anxiety-related behaviors to an equal extent by reducing inflammation, modifying 5-HT receptor expression changes, and inhibiting the ERK pathway. Therefore, MSCs-derived CM may be considered a promising therapy for comorbid anxiety in septic patients.

The Effects of Synbiotic Supplementation on Serum Anti-Inflammatory Factors in the Survivors of Breast Cancer with Lymphedema following a Low Calorie Diet: A Randomized, Double-Blind, Clinical Trial.

BACKGROUND AND AIM: Breast cancer-related lymphedema (BCRL) is a treatment-related inflammatory complication in breast cancer survivors (BCSs). This study was aimed to evaluate the effect of synbiotic supplementation on serum concentrations of IL-10, TGF-ß, VEGF, adiponectin, and edema volume among overweight or obese BCSs with lymphedema following a low-calorie diet (LCD). METHOD: In a randomized double-blind, controlled clinical trial, 88 obese and overweight BCSs women were randomized to synbiotic supplement (n = 44) or placebo (n = 44) groups and both groups followed an LCD for 10 weeks. Pre- and post-intervention comparisons were made regarding the anti-inflammatory markers which included IL-10, TGF-ß, VEGF, adiponectin, edema volume, and anthropometric measurements. Also, the same factors were analyzed to find inter-group disparities. RESULTS: There were no significant differences among participants in the baseline, except for IL-10 and adiponectin. Post-intervention, no significant differences were observed regarding the anti-inflammatory markers, including IL-10, VEGF, adiponectin, and TGF-ß between the groups. After 10 weeks of intervention edema volume significantly decreased in the synbiotic group; additionally, anthropometric measurements (body weight, BMI, body fat percent, and WC) decreased in both groups significantly (P < 0.001 and P < 0.005; respectively). CONCLUSION: Synbiotic supplementation coupled with an LCD in a 10-week intervention had beneficial effects on increasing the serum TGF-ß, IL-10, and adiponectin levels in women with BCRL. It also reduced arm lymphedema volume. Therefore, synbiotic supplementation can be effective in improving health status in BCRL patients.

The Association of EBV and HHV-6 Viral Load with Different NK and CD8+ T Cell Subsets in The Acute Phase of Relapsing-Remitting Multiple Sclerosis.

OBJECTIVE: Epstein-Barr virus (EBV) and Human Herpes virus 6 (HHV-6) are believed to involve in multiple sclerosis (MS) pathogenesis. Natural killer (NK) and CD8+ T cells have essential roles in handling viral infections and their phenotypic and functional properties may be influenced following exposure to viral infections. Here, we investigated the association of NK and CD8+ T cells subpopulations frequency with EBV and HHV-6 viral load in MS patients. MATERIALS AND METHODS: In this case-control study, EBV and HHV-6 viral load were evaluated in plasma of newly diagnosed relapsing-remitting MS (RRMS) patients at relapse phase (n=23), who were not on disease-modifying therapy (DMT), and sex- and age-matched healthy controls (n=19) using real-time polymerase chain reaction (PCR). The frequency of NK and CD8+ T cells subsets were assessed by CD27, CD28, CD45RO, CD56, and CD57 markers using flow cytometry. RESULTS: Despite the increased level of EBV viral load in RRMS patients compared to the control group, there was no statistically significant difference in EBV and HHV-6 copy numbers between the studied groups. In addition, a significant decrease was observed in the percentages of CD56bright CD57- and CD56dim CD57+ CD8low CD45RO- NK cells in RRMS patients in comparison to healthy controls. Analysis of CD8+ T cell subsets showed a substantially high proportion of CD27+ CD28+ CD45RO+ CD57- CD8hi T cells in patients at relapse phase compared to controls. The frequency of NK and T cells subtypes was not associated with EBV and HHV6 plasma viral loads. CONCLUSION: These findings further highlight the variation of NK and CD8+ T cells subsets frequency in clinically active RRMS patients. Since the composition of cells was not associated with EBV and HHV-6 viral load, perhaps other viral infections may be involved in altered NK and CD8+ T cells subpopulation. Larger cohort studies are needed to confirm these results.

Methanolic Extract of Sambucus ebulus Ameliorates Clinical Symptoms in Experimental Type 1 Diabetes through Anti-Inflammatory and Immunomodulatory Actions.

OBJECTIVE: Sambucus ebulus (SE), a famous traditional Iranian medicine, is grown in the north of Iran. As a traditional medicine with anti-inflammatory effects, SE has been utilized against inflammatory joint diseases, insect bites, infectious wounds, edema, and eczema. Type1 diabetes, is an autoimmune disease, characterized by the destruction of pancreatic beta cells by the immune system. For the first time, we investigated the effect of methanolic extract of SE on CD4+, CD8+ and regulatory T cells in experimental type 1 diabetes (T1D). MATERIALS AND METHODS: In this experimental study, fifty-six C57BL\6 mice in 8 groups (G1-G8), were enrolled. Diabetes was induced by a multiple low-dose streptozotocin (MLDS) protocol and mice were daily treated with SE extract at 200 and 400 mg/kg doses, for 35 days. Fasting blood glucose was weekly measured by a glucometer. Islets insulin content was analyzed by immunohistochemistry. Percentage of CD4+, CD8+ and regulatory T cells and cytokines production levels were evaluated by flow cytometer and ELISA, respectively. RESULTS: The clinical symptoms of diabetes were significantly alleviated in G2 group mice which received 400 mg/ kg SE extract. Immunohistochemistry analysis showed that the insulin content of islets increased in G2 group mice. Immunophenotyping analysis indicated that the percentage of CD4+ and CD8+ T cells in G2 group mice was significantly decreased. SE extract significantly increased the percentage of regulatory T cells. The extract in G2 and G4 groups mice significantly decreased IFN-γ and IL-17levels. The extract significantly increased IL-10 in G2 group mice. CONCLUSION: The protective effect of SE extract in MLDS-induced diabetes could be partly due to a decrease of CD4+ and CD8+ T cells and an increase of Treg cells resulting in an inflammation reduction in the pancreatic islets.

RNA Sequencing of CD4+ T Cells in Relapsing-Remitting Multiple Sclerosis Patients at Relapse: Deciphering the Involvement of Novel genes and Pathways.

CD4+ T cells are known as a noteworthy potential modulator of inflammation in multiple sclerosis (MS). In the current study, we investigated the transcriptome profile of CD4+ T cells in patients with relapsing-remitting MS (RRMS) at the relapse phase. We performed RNA sequencing of CD4+ T cells isolated from four relapsing-remitting MS (RRMS) patients at the relapse phase and four age- and sex-matched healthy controls. The edgeR statistical method was employed to determine differentially expressed genes (DEGs). Gene set enrichment analysis was subsequently performed. Applying a physical interaction network, genes with higher degrees were selected as hub genes. A total of 1278 and 1034 genes were defined at significantly higher or lower levels, respectively, in CD4+ T cells of RRMS patients at the relapse phase as compared with healthy controls. The top up- and downregulated genes were JAML and KDM3A. The detected DEGs were remarkable on chromosomes 1 and 2, respectively. The DEGs were mainly enriched in the pathways "regulation of transcription, DNA-templated," "regulation of B cell receptor signaling pathway," "protein phosphorylation," "epidermal growth factor receptor signaling pathway," and "positive regulation of neurogenesis." Moreover, 16 KEGG pathways mostly associated with the immune system and viral infections were enriched. In the constructed physical interaction networks, UBA52 and TP53 were shown to be the most highly ranked hub genes among upregulated and downregulated genes, respectively. By applying global transcriptome profiling of CD4+ T cells, we deciphered the involvement of several novel genes and pathways in MS pathogenesis. The present results must be confirmed by in vivo and in vitro studies.

Exome sequencing reveals novel rare variants in Iranian familial multiple sclerosis: The importance of POLD2 in the disease pathogenesis.

The prevalence of familial multiple sclerosis (FMS) is increasing worldwide which endorses the heritability of the disease. Given that many genome variations are ethnicity-specific and consanguineous marriage could affect genetic diseases, hereditary disease gene analysis among FMS patients from Iran, a country with high rates of parental consanguinity, could be highly effective in finding mutations underlying disease pathogenesis. To examine rare genetic mutations, we selected three Iranian FMS cases with ≥3 MS patients in more than one generation and performed whole exome sequencing. We identified a homozygous rare missense variant in POLD2 (p. Arg141Cys; rs372336011). Molecular dynamics analysis showed reduced polar dehydration energy and conformational changes in POLD2 mutant. Further, we found a heterozygote rare missense variant in NBFP1 (p. Gly487Asp; rs778806175). Our study revealed the possible role of novel rare variants in FMS. Molecular dynamic simulation provided the initial evidence of the structural changes behind POLD2 mutant.

Decreased Serum Levels of Interleukin-4 and Interleukin-21 in New Pemphigus Vulgaris Patients, but Not Chronic Patients With Inactive Disease Compared to Healthy Controls.

BACKGROUND: Pemphigus is a rare group of autoimmune blistering diseases with unknown etiology and unclear pathogenesis. Pemphigus vulgaris (PV) is the most common subtype, and is characterized by ulcerations or flaccid blisters on mucous membranes and on the skin. It is accepted that cytokines have a critical role in the pathogenesis of PV, while their exact roles remain to be elucidated. OBJECTIVES: This study assessed serum levels of interleukin (IL)-4 and IL-21 in different phases of the disease in comparison with healthy controls. METHODS: In a case-control cohort design, serum levels of IL-4 and IL-21 were determined by ELISA in three groups: patients with newly diagnosed PV, patients with chronic, inactive PV (PV in remission), and healthy controls. RESULTS: The study included 88 individuals (58 women and 30 men), including 26 with newly diagnosed PV, 33 with PV in remission, and 29 healthy controls. A significant difference was found among the groups for IL-21 (P = .044), but not for IL-4 (P = .374). Serum levels of IL-4 and IL-21 in newly diagnosed patients were significantly lower than in healthy controls (P = .005 for both), but these cytokine levels in patients with PV in remission were not different from those of controls (P = .343 and P = .221, respectively). Also, no differences in cytokine levels were detected between the newly diagnosed patients and patients with PV in remission. Regardless of disease phase, we detected significantly lower levels of IL-21 in patients than controls (P = .027), but no differences for IL-4 (P = .374). CONCLUSIONS: IL-4 and IL-21 are involved in PV pathogenesis and disease severity. More studies are required to clarify the role of IL-4 and IL-21 in immunopathogenesis and immune response during PV.

The Frequency of CD4+ T Cells in Women with Hashimoto's Thyroiditis.

BACKGROUND: Hashimoto's thyroiditis (HT) is the most prevalent autoimmune disease, and there is no definitive treatment available for this disease. To find the appropriate therapeutic approach, it is necessary to determine the mechanism of this disease. To achieve this purpose, the frequency of CD4+ T cells was evaluated in patients with HT and compared with healthy individuals. METHODS: Twenty-six female patients with HT, aged 20 - 45 years, enrolled in this study. Based on the level of thyroglobulin antibody (anti-TG) and anti-thyroid peroxidase antibody (anti-TPO) in serum of patients with HT, they were divided into two groups. The serum level of anti-TPO was above 100 IU/mL in the group 1 (n = 13), whereas the serum levels of both anti-TPO and anti-TG were above 100 IU/mL in the group 2 (n = 13). Eleven healthy women were considered control group, or group 3. Using flow cytometry, the frequency of T helper (Th)1, Th2, Th17, T regulatory type 1 (Tr1), and LT CD4+IL-4+IL-17+ cells and mean fluorescent intensity (MFI) of their related cytokines were evaluated. RESULTS: The frequency of Th2 cells in the groups 1 (anti-TPO > 100) and 2 (anti-TPO > 100 and anti-TG > 100) were more than control group. Only the difference between groups 3 (healthy control) and 2 was significant (P = 0.022). The frequency of LT CD4+IL-4+IL-17+ cells in the group 1 was significantly more than group 3 (P = 0.027); However, the difference between group 2 and 3 was not significant (P = 0.126). The expression of interferon-gamma (IFN-γ) in the group 2 (P = 0.001) and group 1 (P = 0.001) was significantly higher than group 3. The frequency of Th17, Th1, and Tr1 cells and MFI of IL-17 and IL-10 were not significantly different between the study groups. CONCLUSIONS: In the present study, no significant differences were observed in the frequency of Th17 and Tr1 cells and in MFI of IL-17 and IL-10 in comparison to healthy individuals. Therefore, trying to make a change in the population of these cells probably does not have a significant therapeutic effect. Since Th2 cells and the expression of IFN-γ increased in women with HT, reducing the frequency of Th2 cells or the expression of IFN-γ may be effective in controlling the disease progression. It may be helpful for these patients to prevent the progression of the disease.

Umbilical cord mesenchymal stem cells as well as their released exosomes suppress proliferation of activated PBMCs in multiple sclerosis.

Multiple sclerosis (MS) is a central nervous system (CNS) degenerative disorder which is caused by a targeted autoimmune-mediated attack on myelin proteins. Previously, mesenchymal stem cells were considered as a novel and successful treatment of MS. One of the underlying mechanisms behind their immunomodulatory function is the release of extracellular vesicles, particularly exosomes. In this study, we aimed to evaluate the suppressive efficacy of MSCs and their exosomes on the proliferation of peripheral mononuclear blood cells (PBMC) in relapsing-remitting MS (RRMS) patients and healthy subjects. To do, mesenchymal stem cells were derived from human umbilical cord tissues and used for exosome isolation through ultracentrifugation. Suppressive function of MSCs and MSC-derived exosomes was examined in a coculture with CFSE-labelled PBMCs in vitro. PBMC proliferation of the patients and healthy individuals was measured using flow cytometry. We first demonstrated that proliferation of PBMCs decreased in the presence of MSCs and suppression was more efficient by MSC-derived exosomes, with a minimum alloreaction rate. However, suppression capacity of MSCs and their exosomes significantly decreased during extensive sub-culturing. The present study showed that MSC-derived exosomes as an effective cell-free therapy could prevent proliferation of PBMCs. However, further evaluations are need to move towards a functional approach that can be translated to the clinic.

The inhibitory effect of melatonin on the proliferation of irradiated A549 cell line.

BACKGROUND: Lung cancer is the leading cause of cancer-related deaths worldwide. The high resistance of this type of cancer to radiotherapy and chemotherapy is the greatest challenge for the complete eradication of cancer cells. Although the combination of chemotherapeutic agents has some promising results, severe side effects may limit the received tumor dose. The current study aimed at evaluating the possible synergic effect of melatonin on radiation-induced apoptosis and cell proliferation inhibition. MATERIALS AND METHODS: A549 cells were incubated with melatonin or vehicle and then irradiated with a single dose of 0, 0.5, 2, or 8 Gy X-rays. The cells were incubated with 1 nM of melatonin or vehicle for 1 week and then treated with 1 mM of melatonin or vehicle 1 h before irradiation. Cell proliferation was assessed using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, and apoptosis was assessed using flowcytometry detection of annexin V. RESULTS: Irradiation of the cells with different X-ray doses had no significant impact on MTT results. However, the administration of 1 mM of melatonin 1 h before irradiation significantly reduced the cell proliferation. Nonetheless, there was no significant difference between this treatment group and 1 mM melatonin group. Moreover, the administration of melatonin in combination with irradiation did not show any significant effects on radiation-induced apoptosis. CONCLUSION: The current study results indicated that the treatment of A549 cells with melatonin could suppress cell proliferation, whereas it did not mediate the induction of apoptosis.

Redox Imbalance in CD4+ T Cells of Relapsing-Remitting Multiple Sclerosis Patients.

As a prevalent autoimmune disease of the central nervous system in young adults, multiple sclerosis (MS) is mediated by T cells, particularly CD4+ subsets. Given the evidence that the perturbation in reactive oxygen species (ROS) production has a pivotal role in the onset and progression of MS, its regulation through the antioxidant molecules is too important. Here, we investigated the level of the redox system components in lymphocytes and CD4+ T cells of MS patients. The study was performed on relapsing-remitting MS (RRMS) patients (n = 29) and age- and sex-matched healthy controls (n = 15). Peripheral blood mononuclear cells (PBMCs) were cultured and stimulated by anti-CD3/CD28. The level of ROS, anion superoxide (O2 -), and L-𝛾-glutamyl-Lcysteinylglycine (GSH) was measured by flow cytometry in lymphocytes/CD4+ T cells. The gene expression level of gp91phox, catalase, superoxide dismutase 1/2 (SOD), and nuclear factor-E2-related factor (Nrf2) was also measured by real-time PCR. We found that lymphocytes/CD4+ T cells of RRMS patients at the relapse phase significantly produced higher levels of ROS and O2 - compared to patients at the remission phase (P value < 0.001) and healthy controls (P value < 0.001 and P value < 0.05, respectively). Interestingly, the gene expression level of gp91phox, known as the catalytic subunit of the NADPH oxidase, significantly increased in MS patients at the relapse phase (P value < 0.05). Furthermore, the catalase expression augmented in patients at the acute phase (P value < 0.05), while an increased expression of SOD1 and Nrf2 was found in RRMS patients at relapse and remission phases (P value < 0.05). The increased production of ROS in CD4+ T cells of RRMS patients highlights the importance of amplifying antioxidant components as an efficient approach to ameliorate disease activity in MS patients.

Exogenous Ghrelin Could Not Ameliorate 3,4-methylenedioxymethamphetamine-induced Acute Liver Injury in The Rat: Involved Mechanisms.

MDMA (3,4-methylenedioxymethamphetamine, ecstasy) is often abused by youth as a recreational drug. MDMA abuse is a growing problem in different parts of the world. An important adverse consequence of the drug consumption is hepatotoxicity of different intensities. However, the underlying mechanism of this toxicity has not been completely understood. Ghrelin is a gut hormone with growth hormone stimulatory effect. It expresses in liver, albeit at a much lower level than in stomach, and exerts a hepatoprotective effect. In this study, we investigated hepatotoxicity effect of MDMA alone and its combination with ghrelin as a hepatoprotective agent. MDMA and MDMA+ ghrelin could transiently increase serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) followed by tissue necrosis. However, they could significantly decrease liver tumor necrosis factor-a (TNF-±) in both treatment groups. Unexpectedly, in MDMA treated rats, Bax, Bcl-xl, Bcl-2, Fas, Fas ligand (Fas-L), caspase 8, cytochrome c, caspase 3 gene expression, and DNA fragmentation were nearly unchanged. In addition, apoptosis in MDMA+ ghrelin group was significantly reduced when compared with MDMA treated animals. In all, MDMA could transiently increase serum transaminases and induce tissue necrosis and liver toxicity. Ghrelin, however, could not stop liver enzyme rise and MDMA hepatotoxicity. MDMA hepatotoxicity seems to be mediated via tissue necrosis than apoptotic and inflammatory pathways. Conceivably, ghrelin as an anti-inflammatory and anti-apoptotic agent may not protect hepatocytes against MDMA liver toxicity.

The Influence of Reactive Oxygen Species in the Immune System and Pathogenesis of Multiple Sclerosis.

Multiple roles have been indicated for reactive oxygen species (ROS) in the immune system in recent years. ROS have been extensively studied due to their ability to damage DNA and other subcellular structures. Noticeably, they have been identified as a pivotal second messenger for T-cell receptor signaling and T-cell activation and participate in antigen cross-presentation and chemotaxis. As an agent with direct toxic effects on cells, ROS lead to the initiation of the autoimmune response. Moreover, ROS levels are regulated by antioxidant systems, which include enzymatic and nonenzymatic antioxidants. Enzymatic antioxidants include superoxide dismutase, catalase, glutathione peroxidase, and glutathione reductase. Nonenzymatic antioxidants contain vitamins C, A, and E, glutathione, and thioredoxin. Particularly, cellular antioxidant systems have important functions in maintaining the redox system homeostasis. This review will discuss the significant roles of ROS generation and antioxidant systems under normal conditions, in the immune system, and pathogenesis of multiple sclerosis.

Decreased serum levels of interleukin-17, interleukin-23, TGF-ß in pemphigus vulgaris patients, and their association with disease phase.

The exact pathogenesis of Pemphigus Vulgaris (PV) has remained unclear, but it seems that cytokines play critical roles in this disease. This study aims to assess the serum levels of interleukin (IL)-6, IL-17, IL-23, and TGF-ß in PV patients and compare the results to the healthy controls. Serum levels of IL6, IL-17, IL-23, and TGF-ß were successfully determined by enzyme-linked immunosorbent assay (ELISA) in 27 newly diagnosed PV, 32 patients in remission, and 29 healthy controls. It was shown that the mean serum levels of IL-17, IL-23, and TGF-ß serum are significantly different among the PV patients and healthy controls (P values: <.001, .001, and .003, respectively). It was found that new PV patients have lower serum levels of IL-17, IL-23, and TGF-ß as compared to healthy controls (P values: <.001, <.001, and .003, respectively). Regarding IL-6, no significant difference was observed between the healthy controls and the other two groups of patients. IL-17, IL-23, and TGF-ß are involved in the pathogenesis of PV. However, more studies are required to clarify their exact roles in the immunopathogenesis of PV.

Adipose-Derived Mesenchymal Stem Cells and Conditioned Medium Attenuate the Memory Retrieval Impairment During Sepsis in Rats.

In this study, we hypothesized that sepsis induction impairs memory retrieval in rats while transplanted mesenchymal stem cells (MSCs) and MSC-conditioned medium (MSC-CM) application are capable of attenuating those complications. MSCs were obtained from adipose tissue of rats and at the second culture passage; MSCs and MSC-CM were collected. Rats were randomly divided into four experimental groups: sham, CLP, MSC, and MSC-CM. Sepsis was induced by cecal ligation and puncture (CLP) model in the CLP, MSC, and MSC-CM groups. The MSC group received 1 × 106 MSCs/rat (i.p., 2 h after CLP surgery); the MSC-CM rats received the conditioned medium (CM) from 1 × 106 MSCs intraperitoneally 2 h after sepsis induction. Novel object recognition test, sepsis score, and blood pressure measurement were performed 24 h after the treatments. The right hippocampus was taken for western blot analysis. CLP rats showed a significantly higher sepsis score and systolic blood pressure. They also had a significant increase in the phosphorylated form of CAMKII-α, cleaved caspase 3 and Bax/Bcl2 ratio, and a reduction in c-fos protein in the hippocampus tissue samples compared with the sham group. MSC transplantation and MSC-CM administration significantly decreased the mean sepsis score and prevented sepsis-induced attenuation of blood pressure compared with the CLP rats. Animals in the MSC and MSC-CM groups showed a better memory retrieval, attenuation in phosphorylated form of CAMKII-α, cleaved caspase 3 and Bax/Bcl2 ratio, and an increase in c-fos protein expression compared with the CLP group. It seems that CAMKII and c-fos are inversely involved in regulating memory processes in hippocampus. Phosphorylated form of CaMKII-α overexpression may impair the ability of object recognition. Our findings confirmed that MSC-CM application has more advantages compared with transplanted MSCs and may be offered as a promising therapy for inflammatory diseases such as severe sepsis.