Cytotoxic Cells Research Articles

Targeted therapy of multiple myeloma by IL21-NKG2D CAR-T cells.

NKG2D chimeric antigen receptor (CAR)-modified T cells (NKG2D CAR-T cells) have been reported to be preclinically efficient in several tumors, but little is known whether NKG2D CAR-T cells co-expressing IL21 (IL21-NKG2D CAR-T cells) display greater antitumor activity in multiple myeloma (MM). In this study, the lentivirus has been produced for expression of the IL21 sequence linked to the extracellular NKG2D sequence with the signal peptide linked through the CD8α hinge-transmembrane domain to the 4-1BB molecule fused with the CD3-ζ chain signaling domain, and the engineered IL21-NKG2D CAR-T cells and NKG2D CAR-T cells were constructed. The CAR expression on CAR-T cells was assessed by flow cytometry, and the killing effects of CAR-T cells on MM were assessed by the cytotoxicity assay and ELISA assay. Moreover, xenograft models were also established to evaluate the ability of IL21-NKG2D-CAR-T cells to eliminate MM in vivo. Our results indicated that NKG2D CAR-T cells had dramatic cytotoxicity on MM cells in vitro, and co-expression of IL-21 significantly increased the cytotoxicity of NKG2D CAR-T cells on MM cells. Remarkably, we found that dexamethasone enhanced the cytotoxicity of IL21-NKG2D CAR-T cells on MM cells. Furthermore, IL21-NKG2D CAR-T cells also displayed significant anti-myeloma activity in vivo. In conclusion, IL21-NKG2D CAR-T cells had dramatic cytotoxicity on MM cells in vitro and in vivo, and a system to apply IL21-NKG2D CAR-T cells and low dosage of dexamethasone for the future study of the targeted therapy for MM has been established.

Effect of Acacia concinna Extract on Apoptosis Induction Associated with Endoplasmic Reticulum Stress and Modulated Intracellular Signaling Pathway in Human Colon HCT116 Cancer Cells.

Colorectal cancer (CRC) stands as one of the most prevalent cancer types and among the most frequent causes of cancer-related death globally. Acacia concinna (AC) is a medicinal and edible plant that exhibits a multitude of biological properties, including anticancer properties. This study aimed to investigate the impact of the AC extract on apoptosis induction and the underlying mechanisms associated with this effect in KRAS-mutated human colon HCT116 cells. The effect of AC extract on cell cytotoxicity was evaluated using MTT assay. Nuclear morphological changes were visualized with Hoechst 33342 staining, while mitochondrial membrane potential (MMP) was assessed via JC-1 staining. Flow cytometry was employed for cell cycle analysis, and intracellular ROS levels were determined using DCFH-DA staining. The results showed that HCT116 cells exposed to AC extract showed reduced cell growth and prompted apoptosis, as indicated by an increase in chromatin condensation, apoptotic bodies, the sub-G1 apoptotic cell population, and disrupted MMP. Expression levels of apoptosis mediator proteins determined by Western blot analysis showed an increase in pro-apoptotic proteins (Bak and Bax) while decreasing anti-apoptotic proteins (Bcl-2, Bcl-xL, and Mcl-1), leading to caspase-7 activation and PARP inactivation. AC extract was also found to enhance intracellular reactive oxygen species (ROS) levels and stimulate endoplasmic reticulum (ER) stress. Furthermore, AC extract increases the phosphorylation of ERK1/2, p38, and c-Jun while downregulating PI3K, Akt, β-catenin, and their downstream target proteins. These results demonstrate that AC extract could inhibit cancer cell growth via ROS-induced ER stress associated with apoptosis and regulate the MAPK, PI3K/Akt, and Wnt/β-catenin signaling pathways in HCT116 cells. Therefore, AC extract may be a novel candidate for natural anticancer resources for colon cancer treatment.

Insight into Antiviral Activity of Ag/TiO2 Nanocomposites Against Influenza H1N1 Virus and Its Antiviral Mechanism.

Synthesis and characterization of silver (Ag)/titanium dioxide (TiO2) nanocomposite (ATA) to investigate its antiviral activity against the H1N1 influenza virus and antiviral mechanisms. A water-dispersible ATA was prepared by a photocatalytic reduction process from AgNO3 and TiO2. The characterization of ATA was performed by ultraviolet-visible spectroscopy, X-ray diffraction, high-resolution transmission electron microscopy and energy-dispersive X-ray spectroscopy. The antiviral activities and the antiviral mechanism of ATA were investigated in detail by light microscopy, transmission electron microscopy and biological techniques such as cell cytotoxicity, 50% tissue culture infectious dose detection, western blot and reverse transcription-polymerase chain reaction. These results showed the successful synthesis of ATA nanocomposite with uniform particle size and distribution. It demonstrated the highly efficient antiviral activity of ATA in a dose- and time-dependent manner, as indicated by the reduction of viral titer and the reduction of cytopathic effects caused by viral infection. In the presence of ATA, the structure of the H1N1 influenza virus is directly destroyed and even disintegrated, with the damaged surface membrane proteins and fuzzy contour. It reduces the infection efficiency of influenza by suppressing the activity and expression of hemagglutinin and neuraminidase. The results of mechanistic studies suggested that ATA nanocomposite primarily interferes with virus attachment to viral receptors on the cell surface. Our study suggests that ATA may be a good antiviral candidate against the influenza virus. Compared with AgNPs alone, our synthesized ATA nanocomposites can achieve similar viral inactivation rates using only a much smaller concentration of AgNPs, greatly reducing the amount of AgNPs and their potential side effects. It has great practical value for attaching ATA to the high-efficiency particulate air network in the air purifier, which can kill the virus attached to it and limit its spread.

Identification of PSMD2 as a promising biomarker for pancreatic cancer patients based on comprehensive bioinformatics and in vitro studies

BackgroundPancreatic cancer patients have limited treatment options and extremely poor prognosis. Dysregulations of proteasome 26S subunit, non-ATPases (PSMDs) contribute to the development of various cancers, whereas the significance of PSMDs in pancreatic cancer is poorly understood. In the present study, we intended to explore the therapeutic potential of PSMDs in pancreatic cancer. MethodsBased on TCGA database, the expression of PSMDs was analyzed in pancreatic cancer patients. Multivariate Cox regression and Kaplan–Meier survival analyses were conducted to investigate the prognostic value of PSMDs. The correlations between the expression of PSMD2/14 and immune cell infiltration, immune checkpoint genes’ expression, enrichment of signaling pathways, and the sensitivity of chemotherapies were also evaluated. Knockdown and overexpression experiments were performed to explore the biological function of PSMD2/14. Immunoblotting was conducted to detect the downstream signaling pathway of PSMD2. ResultsMost of the PSMDs, except for PSMD5 and PSMD6, were significantly upregulated in pancreatic cancer tissues. Patients with higher grade tumor had increased mRNA levels of PSMD1/2/5/7/8/11/12/14. Survival and multivariate Cox regression analyses indicated that PSMD2 and PSMD14 were biomarkers of worse prognosis. High expression of PSMD2 and PSMD14 was positively correlated with the levels immune checkpoint genes but not with the infiltration of specific immune cell types. In vitro knockdown of PSMD2, but not PSMD14, increased the apoptosis, gemcitabine’s toxicity and inhibited the growth capacity of MIA cells. Conversely, decreased apoptosis and gemcitabine sensitivity along with accelerated cell proliferation ability were observed in PSMD2-overexpressing PANC-1 cells. Mechanistically, PSMD2 activated the AKT/mTOR signaling pathway, consistent with the findings from KEGG and GSEA analysis. The AKT specific inhibitor MK-2206 exhibited higher cytotoxicity in MIA and PANC-1 cells with high PSMD2 expression. Importantly, MK-2206 largely reversed the oncogenic functions of PSMD2 on the growth and proliferation of PANC-1 cells. ConclusionIn summary, our study provided a comprehensive bioinformatics analysis of PSMDs in pancreatic cancer. We identified that PSMD2 acted as a tumor-promoting protein in pancreatic cancer through the activation of the AKT/mTOR pathway. The overexpression of PSMD2 may be a potential biomarker that predicts the response of pancreatic cancer patients to AKT inhibitor treatments.

Investigating the therapeutic potential of Elemene emulsion injection as an adjuvant for chimeric antigen receptor T cell therapy: Transcriptome analysis and experimental validation

IntroductionElemene emulsion injection (EEI) is prepared from monomers extracted from Chinese medicine. It has been used as an adjuvant anti-tumor medication in clinical settings for over 30 years. Chimeric antigen receptor T cells (CAR-T) are a successful immunotherapy strategy for tumor treatment. This study aimed to investigate the role and potential mechanisms of EEI as an adjuvant to CAR-T cells for tumor treatment. MethodsCCK8 staining was utilized to detect the effect of EEI on tumor cell viability. CD19 CAR-T and Trop2 CAR-T cells were constructed, and EEI was added to the experimental system. Untreated CAR-T cells served as a control group. After co-cultured with tumor cells, the effect of EEI on CAR-T cell function was assessed. Flow cytometry, luciferase reporter gene, IncuCyte, and cytokine assays were used to evaluate the effects of the EEI on CAR-T cell cytotoxicity and phenotype in vitro. RNA sequencing was utilized to analyze the effect of EEI on CAR-T cell gene expression. Network pharmacology was applied to predict potential EEI therapeutic targets for CRS, which were verified via a THP-1 macrophage inflammation model. ResultsEEI enhanced the cytotoxicity of CAR-T cells against tumors. RNA-seq revealed that EEI modulated cytokine activity and Th17 cell differentiation in CAR-T cells. Subsequent experiments confirmed that the proportion of CD4+ T cells in CAR-T cells increased. The secretion of cytokines was greatly reduced under the effect of EEI. In the THP-1 macrophage inflammation model, EEI significantly reduced the levels of cytokines associated with inflammation induced by CAR-T cells. ConclusionsThis research proposed and tested a combination therapy involving EEI and CAR-T cells. The combination of CAR-T cells with EEI for tumor immunotherapy may enhance the clinical efficacy while providing synergistic effects and reducing toxicity. FundingScientific Research Startup Funds for High-level Talents from Beijing University of Chinese Medicine (Grant No. 9011451310032)

Green approach of cobalt sulfide nanoparticles from novel red stigma of Crocus sativus and multifaceted biomedical advancement

The present study was based on green protocols for synthesizing cobalt sulfide nanoparticles using saffron stigma flower extract (Crocus sativus), as saffron is considered a potent traditional medicine. Furthermore, the novel cobalt sulfide (CoS) nanoparticles prepared by the green approach for the first time with saffron improved showed cytotoxicity and antibacterial activity, as well as significant antitumor activity of Hela, A549, and MCF 7 cells viz in vitro studies. The saffron-cobalt sulfide nanoparticles underwent characterization using various techniques which include the X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FTIR), Advances in scanning electron microscopy (SEM), and high-resolution transmission electron microscopy (HR-TEM), and the energy dispersive X-ray analysis spectroscopy. Both XRD, and FTIR confirmed the synthesis of pure cobalt sulfide nanoparticles. The resulting saffron-based cobalt sulfide nanoparticles exhibited a nanosphere structure, as confirmed by HRTEM, indicating the formation of nanosphere and microstructures. In vitro studies were conducted to assess the cytotoxicity and cell viability of Hela cells, A549, and MCF-7 cells. And, antibacterial studies were performed against various bacterial strains using different concentrations of cobalt sulfide nanoparticles derived from the red stigma of saffron (Crocus sativus) only the red stigma extract. The MTT assay analysis, cell viability, and morphological studies demonstrated the cytotoxic studies of the synthesized cobalt sulfide nanoparticles on cancer cells. The antibacterial test of the nanoparticles was also evaluated, revealing significant zones of inhibition (mm) against pathogens and cobalt sulfide nanoparticles using saffron extract, showcasing their potent cytotoxic, antibacterial, and antitumor activities with promising biomedical applications.

Novel tri-specific T-cell engager targeting IL-13Rα2 and EGFRvIII provides long-term survival in heterogeneous GBM challenge and promotes antitumor cytotoxicity with patient immune cells

BackgroundGlioblastoma multiforme (GBM) is known for its high antigenic heterogeneity, which undermines the effectiveness of monospecific immunotherapies. Multivalent immunotherapeutic strategies that target multiple tumor antigens simultaneously could enhance clinical outcomes...

Atopic Dermatitis Immune Dysregulation as Dengue Predisposing Factor.

The immune response is important in dengue's clinical manifestation, and the immune dysregulation in Atopic Dermatitis (AD) can permit immune evasion by viruses. There have been many studies describing the immune response in AD and the pathomechanism of dengue, but AD as a predisposing factor for dengue and its severity have not been much discussed. This review investigates how immune dysregulation in AD may be a predisposing factor for Dengue and its severe outcomes. We conducted a comprehensive analysis of studies from the past decade, focusing on dendritic cells (DCs), macrophages, mast cells (MCs), Innate Lymphoid Cell 2 (ILC-2), Natural Killer (NK) cells, interferon (IFN), interleukin (IL) 4, IL-13, and T helper (Th) 2 in AD patients with healthy subject as a comparison, using databases PubMed, Science Direct, and Google Scholar. We got 44 articles that met inclusion criteria. From those articles, we resumed that moderate and severe AD patients' immune profiles showed increased DC, MCs, M2 macrophage, NK cells, and ILC2 in the lesional and non-lesional skin, decreased DC and NK cells in peripheral blood, alteration cytotoxicity of NK cells,Th2-skewed adaptive immune response in lesional and non-lesional skin, and peripheral blood. Increased DC, M2 macrophage, and MCs provide target cells for Dengue virus (DENV) replication. Alteration cytotoxicity of NK cells, ILC2, and Th2 skewed immune response facilitated immune evasion by DENV. The innate and adaptive immune dysregulation in moderate and severe AD provides DENV target cells and facilitates virus immune evasion that can be a predisposing factor for dengue and severe dengue. Further research is recommended to clarify the association between AD and the incidence of dengue and severe dengue because this can be a consideration in determining the prognosis and management of Dengue.

Potential treatment approaches for malignant peritoneal mesothelioma: in vivo and in vitro experimental study of natural killer cell immunotherapy.

Malignant peritoneal mesothelioma (MPM) is a rare primary malignant tumor with an extremely poor prognosis that currently lacks effective treatment options. This study investigated the in vitro and in vivo efficacy of natural killer (NK) cells for treatment of MPM. An in vitro study was conducted to assess the cytotoxicity of NK cells from umbilical cord blood to MPM cells with the use of a high-content imaging analysis system, the Cell Counting Kit-8 assay, and Wright-Giemsa staining. The level of NK cell effector molecule expression was detected by flow cytometry and enzyme-linked immunosorbent assays. The ability of NK cells to kill MPM cells was determined based on live cell imaging, transmission electron microscopy, and scanning electron microscopy. An in vivo study was conducted to assess the efficacy and safety of NK cell therapy based on the experimental peritoneal cancer index, small animal magnetic resonance imaging, and conventional histopathologic, cytologic, and hematologic studies. NK cells effectively killed MPM cells through the release of effector molecules (granzyme B, perforin, interferon-γ, and tumor necrosis factor-α) in a dose- and density-dependent manner. The NK cell killing process potentially involved four dynamic steps: chemotaxis; hitting; adhesion; and penetration. NK cells significantly reduced the tumor burden, diminished ascites production, and extended survival with no significant hematologic toxicity or organ damage in NOG mice. NK cell immunotherapy inhibited proliferation of MPM cells in vitro and in vivo with a good safety profile.

Metabolic programs drive function of therapeutic NK cells in hypoxic tumor environments.

Limited oxygen (hypoxia) in solid tumors poses a challenge to successful immunotherapy with natural killer (NK) cells. NK cells have impaired cytotoxicity when cultured in hypoxia (1% oxygen) but not physiologic (>5%) or atmospheric oxygen (20%). We found that changes to cytotoxicity were regulated at the transcriptional level and accompanied by metabolic dysregulation. Dosing with interleukin-15 (IL-15) enhanced NK cell cytotoxicity in hypoxia, but preactivation with feeder cells bearing IL-21 and 4-1BBL was even better. Preactivation resulted in less perturbed metabolism in hypoxia; greater resistance to oxidative stress; and no hypoxia-induced loss of transcription factors (T-bet and Eomes), activating receptors, adhesion molecules (CD2), and cytotoxic proteins (TRAIL and FasL). There remained a deficit in CD122/IL-2Rβ when exposed to hypoxia, which affected IL-15 signaling. However, tri-specific killer engager molecules that deliver IL-15 in the context of anti-CD16/FcγRIII were able to bypass this deficit, enhancing cytotoxicity of both fresh and preactivated NK cells in hypoxia.

Circular RNA circ_0061183 regulates microglial polarization induced by airborne ultrafine particles in HMC3 cells via sponging miR-98-5p

Airborne ultrafine particulate matter (PM0.1) can enter the brain, induce microglia activation, and promote the development of Alzheimer’s disease (AD). Circular RNAs (circRNAs) are also involved in AD pathogenesis. However, the role of AD-related circRNAs in PM0.1-induced microglia activation remains unclear. Therefore, we explore cytotoxicity, microglia activation, and AD-associated circRNA expression in human microglial HMC3 cells treated with PM0.1, and further examined circRNA expression in mice and cognitively impaired individuals. The results revealed that PM0.1 exposure decreased cell viability, increased lactate dehydrogenase activity, caused microglia activation, elevated microglial M1 maker expression, downregulated microglial M2 maker expression, and reduced AD-related circ_0061183 expression in vitro. Functionally, circ_0061183 silencing enhanced microglia activation and microglial M1 polarization, but inhibited microglial M2 polarization. Mechanistically, circ_0061183 can bind to miR-98-5p to co-regulate M2 microglial-related IL10 expression, which may affect transforming growth factor-β signaling to regulate PM0.1-inhibited microglial M2 polarization. Moreover, circ_0061183 downregulation was observed in the brain of PM2.5-exposed mice and AD mice and in the blood of cognitively impaired individuals. Furthermore, circ_0061183 was positively related to mini–mental state examination scores and amyloid-β42 peptide expression in elderly individuals. Overall, the current work offers epigenetic insights into the regulatory mechanisms of circRNAs on microglial activation caused by environmental pollutants.

Discovery of a new lead molecule to develop a novel class of human factor XIIa inhibitors.

Factor XIIa (FXIIa) is a plasma serine protease within the contact activation pathway. Inhibiting FXIIa could offer a viable therapeutic approach for achieving effective and safer anticoagulation without the bleeding risks that accompany the use of existing anticoagulants. Therefore, we investigated the anticoagulant properties of an amidine-containing molecule (inhibitor 1) to identify a potential lead molecule for subsequent development of FXIIa inhibitors. Results indicated that inhibitor 1 primarily inhibits human FXIIa with an IC50 value of ~30 µM. The inhibitor demonstrated variable selectivity against thrombin, factor IXa, factor Xa, factor XIa, and activated protein C. Michaelis-Menten kinetics indicated that the molecule is an active site inhibitor of FXIIa. Molecular modeling studies revealed that the molecule recognizes residues His57, Asp189, and Ala190 in FXIIa's active site. The inhibitor selectively and concentration-dependently prolonged the clotting time of human plasma under activated partial thromboplastin time assay conditions. The inhibitor did not exhibit significant cytotoxicity in human HEK293 cells and the in silico pharmacokinetics and toxicology data were comparable to known anticoagulants. This study introduces inhibitor 1 as a lead platform for further development as an anticoagulant to provide a more effective and safer approach to preventing and treating thromboembolic diseases.

ICOS-expressing CAR-T cells mediate durable eradication of triple-negative breast cancer and metastasis

BackgroundThe failure of conventional therapies and the propensity for recurrence and metastasis make triple-negative breast cancer (TNBC) a formidable challenge with grim prognoses and diminished survival rates. Immunotherapy, including immune...

Asciminib stands out as the superior tyrosine kinase inhibitor to combine with anti-CD20 monoclonal antibodies for the treatment of CD20+ Philadelphia-positive B-cell precursor acute lymphoblastic leukemia in preclinical models.

Philadelphia chromosome-positive B-cell precursor acute lymphoblastic leukemia (Ph+ BCP-ALL) is a high-risk subtype of acute lymphoblastic leukemia characterized by the presence of the BCR::ABL1 fusion gene. Tyrosine kinase inhibitors (TKI) combined with chemotherapy are established as the first-line treatment. Additionally, rituximab, an anti-CD20 monoclonal antibody is administered to adult BCP-ALL patients with ≥20% CD20+ blasts. In this study, we observed a marked prevalence of CD20 expression in patients diagnosed with Ph+ BCP-ALL, indicating a potential widespread clinical application of rituximab in combination with TKI. Consequently, we examined the influence of TKI on the antitumor effectiveness of anti-CD20 monoclonal antibodies by evaluating levels of CD20 on the cell surface and conducting in vitro functional assays. All tested TKI were found to uniformly downregulate CD20 on leukemic cells, diminishing the efficacy of rituximab-mediated complement- dependent cytotoxicity. Interestingly, these TKI displayed varied effects on natural killer (NK) cell-mediated antibody- dependent cytotoxicity and macrophage phagocytic function. While asciminib demonstrated no inhibition of effector cell functions, dasatinib notably suppressed the anti-CD20-monoclonal antibody-mediated NK cell cytotoxicity and macrophage phagocytosis of BCP-ALL cells. Dasatinib and ponatinib also decreased NK cell degranulation in vitro. Importantly, oral administration of dasatinib, but not asciminib, compromised NK cell activity in patients' blood, as determined by an ex vivo degranulation assay. Our results indicate that asciminib might be preferred over other TKI for combination therapy with anti-CD20 monoclonal antibodies.

A Novel Combinatorial Regimen Using Sorafenib and Uttroside B, A US FDA-designated 'Orphan Drug', for the Treatment of Hepatocellular Carcinoma.

Sorafenib (Sor) is the first-line treatment option in clinics for treating advanced unresectable hepatocellular carcinoma (HCC). However, acquired chemoresistance and adverse side effects associated with Sor monotherapy limit its clinical benefits. We have previously reported the exceptional anti-HCC potential of uttroside B (Utt-B), a furostanol saponin isolated in our lab from Solanum nigrum Linn. leaves. The current study has evaluated the supremacy of a combinatorial regimen of Sor and Utt-B over Sor monotherapy. MTT assay was used for In vitro cytotoxicity studies. A clonogenic assay was conducted to assess the anti-proliferative effect of the combination. Annexin V/PI staining, confocal microscopy, FACS cell cycle analysis, and Western blotting experiments were performed to validate the pro-apoptotic potential of the combination in HepG2 and Huh7 cell lines. Pharmacological safety evaluation was performed in Swiss albino mice. Our results indicate that Utt-B augments Sor-induced cytotoxicity in HepG2 and Huh7 cells. The combination inhibits the proliferation of liver cancer cells by inducing apoptosis through activation of the caspases 7 and 3, leading to PARP cleavage. Furthermore, the combination does not induce any acute toxicity in vivo, even at a dose five times that of the effective therapeutic dose. Our results highlight the potential of Utt-B as an effective chemosensitizer, which can augment the efficacy of Sor against HCC and circumvent Sor-induced toxic side effects. Moreover, this is the first and only report to date on the chemosensitizing potential of Utt-B and the only report that demonstrates the therapeutic efficacy and pharmacological safety of a novel combinatorial regimen involving Utt-B and Sor for combating HCC.

Biotoxicity Evaluation of Biodegradable Polymeric Particles: Exploring the Possible Adverse Impacts on Biological Systems

ABSTRACTThis study aimed to investigate the impact of plastic particles on cell viability and tissue toxicity. The cells were subjected to incubation with plastic particles immobilized in agar gels, resulting in minimal cell death and cytotoxicity. Furthermore, direct exposure of various cell types to suspended plastic particles showed negligible effects on cell viability, with no observed lysis or growth inhibition. Staining techniques revealed minimal plastic particle‐induced cell death, with the majority of cells remaining viable. However, histological evaluations of mouse tissues demonstrated that the nondegradable low‐density poly(ethylene) (LDPE) groups exhibited severe irritation, characterized by an excessive presence of macrophages, neutrophils, fibrosis, fat infiltration, and increased blood vessel formation in subcutaneous tissue. In contrast, biodegradable neat poly(butylene succinate) (KRICT‐PBS) and cellulose nanocrystals/PBS nanocomposite (CNC‐PBS) groups induced minimal toxicity. Interestingly, the intravenous injection of KRICT‐PBS and CNC‐PBS degradate solutions in mice exhibited mild toxicity, suggesting no significant damage to normal kidney and liver function.

Integrated single-cell transcriptome and TCR profiles of hepatocellular carcinoma highlight the convergence on interferon signaling during immunotherapy

BackgroundDespite the success of immune checkpoint inhibitor (ICI)-based combination therapies in hepatocellular carcinoma (HCC), its effectiveness remains confined to a subset of patients. The development of reliable, predictive markers is...

Evaluation of the immunomodulatory effects of a novel synbiotic made of combined use of probiotic-prebiotic-Chinese traditional herbs

BackgroundRecent studies have discussed the preferred activity of synbiotics, which are a combination of probiotics and prebiotics. This study formulates a novel synbiotic that includes Chinese traditional herbs and evaluates its immunomodulatory function. PurposeThis study aims to systematically investigate the effects of synbiotics on immune function using a cyclophosphamide-induced immunosuppressed mouse model. Study DesignA literature-based approach was used to selectively identify medicinal herbs with immunomodulatory properties. Commercially available probiotics were used to investigate the immunomodulatory functions and potential mechanisms associated with the combination of Chinese medicinal herbs and prebiotics as carbon substrates. MethodsThe immunomodulatory effect of the synbiotic will be assessed by measuring indicators such as body weight, organ index, immunoglobulins, cytokines, splenic lymphocyte proliferation status, NK cell cytotoxicity and gut microbiota. ResultsOur results demonstrated that the synbiotic significantly restored the decreased thymus index and the increased spleen index induced by immune deficiency in mice. Among them, the formula of the synbiotics significantly improves the thymus organ index of immune organs from 0.068 to 0.126, which is also significantly superior compared to other single components. Additionally, it significantly enhanced the proliferative capacity of splenic lymphocytes and the cytotoxic activity of natural killer (NK) cells in immune-deficient mice. Furthermore, the synbiotics enhanced the richness and diversity of the gut microbiota, rectifying the dysbiosis in gut microbial composition caused by immune deficiency. It decreased the relative abundance of Firmicutes from 67.41% to 44.06%, while increasing the relative abundance of Bacteroidota from 30.42% to 53.77%. It restored the disrupted gut microbial structure induced by cyclophosphamide damage, thereby ameliorating the disrupted intestinal microbial community associated with immune deficiency. ConclusionThe study demonstrated that the synbiotic was capable of modulating immune function, as evidenced by an increase in organ index, cellular immunity, and the ability to modulate intestinal flora disorders. Furthermore, the synbiotic demonstrated superior performance compared to other single components.

IL-17 Producing T to Foxp3+CD4+ Regulatory T Cell Ratio as a Diagnostic and Prognostic Marker in Women With Recurrent Pregnancy Loss and Its Implications for Intravenous Immunoglobulin Therapy.

The imbalance in the Th17/Regulatory T (Treg) cell ratio is associated with recurrent pregnancy loss (RPL). This study aimed to determine a cut-off for the Th17/Treg cell ratio to predict pregnancy outcomes in RPL and evaluate the effectiveness of intravenous immunoglobulin (IVIG) based on this cut-off value. This retrospective cohort study included 49 idiopathic RPL and 75 controls. The subgroups of IL-17+ T cell to Foxp3+ T cell ratios in peripheral blood were measured using flow cytometry. The cut-off values of Th17/Treg cell ratios were determined by the ROC curve to distinguish between RPL and controls. The IVIG treatment effectiveness in pregnancy outcome was compared between high- and low-ratio groups. Pearson correlation assessed the Th17/Treg cell ratio's relationship with NK cell cytotoxicity (NKC), NK cell percentage, and Th1/Th2 cell ratio. Using the ROC curve, we identified six Th17/Treg cell ratio markers with diagnostic value, and the following two, CD3+IL-17+ T cell/CD3+Foxp3high T cell ratio (sensitivity at 97%) and CD4+IL-17+ T cell/CD3+Foxp3high T cell ratio (specificity at 93.61%), showed the highest statistical significance in diagnosing idiopathic RPL. Among the six diagnostic markers, in terms of predicting pregnancy outcomes with IVIG treatment, CD3+IL-17+ T cell/CD4+Foxp3+ T cell ratio was the most valuable prognostic marker. In RPL women with high CD3+IL-17+ T cell/CD4+Foxp3+ T cell ratio (≥ 1.096), the live birth rate (LBR) was improved with IVIG treatment. (IVIG treatment, 78.57%vs. no IVIG, 28.57%, p = 0.026). On the other hand, RPL women with low CD3+IL-17+ T cell/CD4+Foxp3+ T cell ratio did not demonstrate the effectiveness of IVIG (LBRs with IVIG treatment, 50.00%vs. no IVIG, 84.62%, p = 0.219). In a correlation study, the CD3+IL-17+ T cell/CD4+Foxp3+ T cell ratio was an independent prognostic marker, showing no correlation with NKC, NK cell percentage, and Th1/Th2 cell ratio. The CD3+IL-17+ T/CD4+Foxp3+ T cell ratio may serve as a valuable marker for understanding the pathogenesis of RPL, predicting pregnancy outcomes, and selecting candidates for immunotherapy. Our study demonstrates that IVIG treatment can significantly improve LBR in women with a high CD3+IL-17+ T/CD4+Foxp3+ T ratio, offering a promising therapeutic approach for this challenging condition.

PEDV evades MHC-I-related immunity through nsp1-mediated NLRC5 translation inhibition.

Major histocompatibility complex class I (MHC-I) plays crucial roles against viral infections not only by initiating CD8+ T cell immunity but also by modulating natural killer (NK) cell cytotoxicity. Understanding how viruses precisely regulate MHC-I to optimize their infection is important; however, the manipulation of MHC-I molecules by porcine epidemic diarrhea virus (PEDV) remains unclear. In this study, we demonstrate that PEDV infection promotes the transcription of NLRC5, a key transactivator of MHC-I, in several porcine cell lines and in vivo. Paradoxically, no increase in MHC-I expression is observed after PEDV infection both in vitro and in vivo. Mechanistic studies revealed that PEDV infection inhibits the translation of PEDV-elicited NLRC5 mRNA and the expression of downstream MHC-I proteins, without affecting the expression of physiological NLRC5 and MHC-I proteins. Through viral protein screening, we identified PEDV nonstructural protein 1 (nsp1) as the critical antagonist that inhibits NLRC5-mediated upregulation of MHC-I, and the nsp1's inhibitory effect on MHC-I requires the motif of 15 amino acids at its C-terminus. Notably, our results revealed that the cytotoxic ability of NK cells against PEDV-infected cells is similar to that against healthy cells. Collectively, our findings uncover an immune evasion mechanism by which PEDV-infected cells masquerade as healthy cells to evade NK and T cell immunity. This is achieved by targeting NLRC5, a key MHC-I transcriptional regulator, via nsp1.IMPORTANCEPorcine epidemic diarrhea virus (PEDV) is a highly contagious enteric coronavirus that inflicts substantial financial losses on the swine industry. Major histocompatibility complex class I (MHC-I) is a critical factor influencing both CD8+ T cell and natural killer (NK) cell immunity. However, how PEDV manipulates MHC-I expression to optimize its infection process remains largely unknown. In this study, we demonstrate that PEDV's nonstructural protein 1 (nsp1) inhibits virus-mediated induction of MHC-I expression by directly targeting NLRC5, a key MHC-I transactivator. Intriguingly, nsp1 does not reduce physiological NLRC5 and MHC-I expression. This selective inhibition of virus-elicited NLRC5 mRNA translation allows PEDV-infected cells to masquerade as healthy cells, thereby evading CD8+ T cell and NK cell cytotoxicity. Our findings provide unique insights into the mechanisms by which PEDV evades CD8+ T cell and NK cell immunity.