Spleen Lymphocyte Proliferation Research Articles

Immunomodulatory Effect of Phage Depolymerase Dep_kpv74 with Therapeutic Potential Against K2-Hypervirulent Klebsiella pneumoniae.

Background: The emergence of multidrug-resistant hypervirulent Klebsiella pneumoniae (hvKp) has made it difficult to treat and control infections caused by this bacterium. Previously, the therapeutic effectiveness of phage-encoded depolymerase Dep_kpv74 in a mouse model of K. pneumoniae-induced thigh soft tissue infection was reported. In this study, the effect of Dep_kpv74 on blood parameters in mice, the proliferation and subpopulation composition of spleen lymphocytes, and the activity and stability of the enzyme at different pH and temperatures were further explored. Results: The stability tests showed that Dep_kpv74 remained active in the temperature range from 8 °C to 55 °C. The optimal pH value for maintaining the activity of Dep_kpv74 ranged from 5.0 to 9.0. The depolymerase was detected in the blood, spleen, and lungs of mice 10 min after intraperitoneal administration, reaching maximum activity values after 1-3 h and maintaining activity a day after administration. The introduction of Dep_kpv74 at the therapeutic dose (10 μg/mouse) or at a 10-fold higher dose did not lead to reliable changes in bloodstream cell content compared with the reference values of intact mice. The biochemical results of the studies indicated that Dep_kpv74 did not exert any toxic effects on liver and kidney functions. The results of the analysis of lymphocyte proliferative activity demonstrated that Dep_kpv74 depolymerase has a mild immunomodulatory effect. Conclusions: Thus, the results of this study provide one more confirmation that depolymerase Dep_kpv74 is a potential candidate for the treatment of infections caused by hvKp expressing K2 capsular polysaccharides.

Preclinical testing of immunotoxicity for animal-derived dermal materials

Animal derived materials have been widely used in biomedical products owing to their good biocompatibility and appropriate mechanical properties. It is crucial to evaluate the immunotoxicity of such materials in preclinical testing to prevent severe immune responses in patients. In this study, a pipeline of immunotoxicity tests was established to evaluate animal-derived materials before de-cellularization and final products. This pipeline contains a serial of animal tests on BALB/c mice and an in vitro quantification test for a-Gal antigen. It is well-recognized that the interaction between materials and patients profoundly alters immune responses, thus, a comprehensive dataset including body weight, immune organ coefficient, histopathology, peripheral hematology, serum immunoglobulin level, spleen lymphocyte proliferation rate, and their subpopulation was created and analyzed using the SPSS tool. These results clearly suggested that the de-cellularized materials possessed better biocompatibility, in addition, the a-Gal antigen content was effectively decreased by 96.0 ​% after de-cellularization. Thus, this study confirmed that this multi-step enzymatic de-cellularization treatment is a potent method to reduce the immunotoxicity of animal-derived biomaterials. Moreover, the experimental pipeline will likely be transferable to other biomedical materials and products.

Potential of Pandan anggur (Sararanga Sinuosa Hemsley) as immunomodulator

Background: Pandan Anggur plant is an endemic plant of the Papua archipelago in Indonesia, and the fruit called Pandan Anggur Fruit (PAF) is freshly consumed or prepared as a juice. Based on previous studies on the phytochemical content of PAF, the ethanol and water extract contain flavonoids, which act as immunomodulators, to regulate the response of immunity. Objectives: This study aimed to examine the potential of PAF as an immunomodulator in malnutrition cases, by evaluating six parameters which are phagocytosis capacity and nitric oxide (NO) production of peritoneal macrophage, spleen lymphocyte proliferation, IFN-γ and IL-4 in lymphocyte culture, and intestinal sIgA levels. By proving the potential of PAF as an immunomodulator, this research will lead to significant development of functional food products of Pandan Anggur. Methods: A total of 35 Sprague Dawley rats were used as animal models divided into seven different groups named standard group (healthy standard), malnourished (negative control), malnourished rats given Imboost Force (positive control), as well as PAF ethanol extract doses of I, II, III, and PAF water extract. The treatment lasted for seven weeks in total and the measurement of immunomodulator parameters was carried out at the end of the treatment period. The immunomodulator parameters included phagocytosis capacity and nitric oxide (NO) production of peritoneal macrophage, spleen lymphocyte proliferation, IFN-γ and IL-4 in lymphocyte culture, and intestinal sIgA levels. Result: The results indicated that the administration of PAF ethanol extract at a dose of 3.15 mg/mL provided an optimum immunostimulant effect, by increasing the macrophage phagocytosis capacity, nitric oxide, lymphocyte proliferation, IL-4, and sIgA levels in rat intestine. However, increasing the dose of extract did not produce a better effect, but led to suppression of immunity. Conclusion: PAF extract at a dose of 3.15 mg/mL provides an immunostimulant effect. Keywords: In Vivo, IL-4, IF-γ, sIgA, lymphocyte proliferation, macrophage, Pandan Anggur Fruit

Alkaloids in Uncaria rhynchophylla improves AD pathology by restraining CD4+ T cell-mediated neuroinflammation via inhibition of glycolysis in APP/PS1 mice

Ethnopharmacological relevanceUncaria rhynchophylla (Miq.) Miq.ex Havil. was a classical medicinal plant exhibiting the properties of extinguishing wind, arresting convulsions, clearing heat and pacifying the liver. Clinically, it could be utilized for the treatment of central nervous system-related diseases, such as Alzheimer's disease. U. rhynchophylla (UR) and its major ingredient alkaloid compounds (URA) have been proved to exert significant neuroprotective effects. However, the potential mechanism aren't fully understood. Aim of the studyThis study systematically examined the therapeutic effects of URA on AD pathology in APP-PS1 mice, and revealed the potential mechanism of action. Materials and methodsThe cognitive ability was evaluated by morris water maze test in APP-PS1 mice. The H&E staining was used to observe the tissue pathological changes. The ELISA kits were used to detect the level of inflammatory factors. The flow cytometry was used to analyze the percentage of CD4+ effector T cells (Teffs) in spleen. The immunofluorescent staining was performed to count the Teffs and microglia in brain. The protein expression was analyzed by western blot. In vitro, the lymphocyte proliferation induced by ConA was performed by CCK-8 kits. The IFN-γ, IL-17, and TNF-α production were detected by ELISA kits. The effects of URA on glycolysis and the involvement of PI3K/Akt/mTOR signaling pathway was analyzed by Lactic Acid assay kit and western blot in ConA-induced naive T cell. ResultsURA treatment improved AD pathology effectively as demonstrated by enhanced cognitive ability, decreased Aβ deposit and Tau phosphorylation, as well as reduced neuron apoptosis. Also, the neuroinflammation was significantly alleviated as evidenced by decreased IFN-γ, IL-17 and increased IL-10, TGF-β. Notably, URA treatment down-regulated the percentage of Teffs (Th1 and Th17) in spleen, and reduced the infiltration of Teffs and microglia in brain. Meanwhile, the Treg cell was up-regulated both in spleen and brain. In vitro, URA was capable of attenuating the spleen lymphocyte proliferation and release of inflammatory factors provoked by ConA. Interestingly, glycolysis was inhibited by URA treatment as evidenced by the decrease in Lactic Acid production and expression of HK2 and GLUT1 via regulating PI3K/Akt/mTOR signaling pathway in ConA-induced naive T cell. ConclusionThis study proved that URA could improve AD pathology which was possibly attributable to the restraints of CD4+ T cell mediated neuroinflammation via inhibiting glycolysis.

Preparation Optimization and Immunological Activity Studies of Portulaca oleracea L. Polysaccharides Liposomes.

This study combines traditional Chinese medicine polysaccharides with nanomaterials to enhance drug bioavailability and immunological activity. The study of polysaccharide preparation, structure identification, pharmacological activity, and mechanism of action is deepening, but the research combined with the new drug delivery system is relatively weak, so the application of polysaccharides is still facing great limitations. In order to prolong the action time of polysaccharides and improve their bioavailability, liposome has become the most promising delivery carrier. The purpose of this study was to optimize the preparation process of Portulaca oleracea L. polysaccharides liposomes (POL-PL) and evaluate the immunoactivity in vitro. POL-PL was prepared by reverse evaporation, and the preparation process was optimized using the response surface methodology. The characteristic analysis of POL-PL was detected by the indicators including morphology, particle size, zeta potential, encapsulation efficiency, release, and stability. The effects of POL-PL on the proliferation and immunological activity of mouse spleen lymphocytes and RAW264.7 cells were evaluated in vitro. POL-PL is highly homogeneous in morphology and particle size, and its sustained release improves the bioavailability of Portulaca oleracea L. polysaccharides (POL-P). Moreover, POL-PL treatment significantly enhanced the proliferation and phagocytic activity of RAW264.7 cells and increased the secretion of IL-6, TNF-α, IL-1β, and NO. This study suggested that POL-PL were prepared successfully by reverse evaporation method, and POL-PL had immunoenhancing activity in vitro. The results provided a theoretical basis for further application of POL-PL.

Bioinformatic features and immunological response of recombinant antigen CTLA4-IgV-EgG1Y162 against Echinococcus granulosus.

Cystic echinococcosis (CE) is a zoonotic disease caused by the infection of Echinococcus granulosus (E. granulosus) larva. Currently, blocking the pathogenic cycle chain through immunoprophylaxis has become the main research direction. EgG1Y162 protein has good antigenicity and immunogenicity and is therefore a good candidate molecule for E. granulosus vaccine. Mature T cells express CTLA-4 on their surface, and its extracellular IgV region binds efficiently to the B7 molecules on antigen-presenting cells to deliver negative signals. We designed and prepared a recombinant vaccine by fusing CTLA-4IgV to the EgG1Y162 protein to exploit its binding properties. Bioinformatic methods were used to analyze the structure and epitopes of the proposed recombinant vaccine. The placement of 16 amino acids (GTDDDDKAMADIGSEF) between the CTLA-4IgV and EgG1Y162 using the skeleton structure of pET30a plasmid did not affect the correct folding of the proteins. When the recombinant proteins were co-cultured with bone marrow-induced dendritic cells (DC), the protein CTLA-4IgV-EgG1Y162 promoted its binding to DC and increased the percentage of DC maturation compared with protein EgG1Y162 in vitro and in vivo. Compared to EgG1Y162, CTLA-4IgV-EgG1Y162 promoted the proliferation of lymphocytes in spleen and the release of interferon (IFN)-γ and interleukin (IL)-4 by those lymphocytes in vitro, while it also promoted the release of protective antibodies in the serum of immunized mice in vivo. These findings indicated that the designed recombinant vaccine, CTLA-4IgV-EgG1Y162, can provide new ideas for the optimization and improvement of vaccines against E. granulosus.

A Unique Combination of Mn2+ and Aluminum Adjuvant Acted the Synergistic Effect.

The development of combinatorial adjuvants is a promising strategy to boost vaccination efficiency. Accumulating evidence indicates that manganese exerts strong immunocompetence and will become an enormous potential adjuvant. Here, we described a novel combination of Mn2+ plus aluminum hydroxide (AH) adjuvant that significantly exhibited the synergistic immune effect. Methodology. Initially, IsdB3 proteins as the immune-dominant fragment of IsdB proteins derived from Staphylococcus aureus (S. aureus) were prepared. IsdB3 proteins were identified by western blotting. Furthermore, we immunized C57/B6 mice with IsdB3 proteins plus Mn2+ and AH adjuvant. After the second immunization, the proliferation of lymphocytes was measured by the cell counting kit-8 (CCK-8) and the level of IFN-γ, IL-4, IL-10, and IL-17 cytokine from spleen lymphocytes in mice and generation of the antibodies against IsdB3 in serum was detected with ELISA, and the protective immune response was assessed through S. aureus challenge. IsdB3 proteins plus Mn2+ and AH obviously stimulated the proliferation of spleen lymphocytes and increased the secretion of IFN-γ, IL-4, IL-10, and IL-17 cytokine in mice, markedly enhanced the generation of the antibodies against IsdB3 in serum, observably decreased bacterial load in organs, and greatly improved the survival rate of mice. These data showed that the combination of Mn2+ and AH significantly acted a synergistic effect, reinforced the immunogenicity of IsdB3, and offered a new strategy to increase vaccine efficiency.

Studies of the Immunomodulatory Activity of Polysaccharides from the Stem of Cynomorium songaricum Based on Intestinal Microbial Analysis.

Polysaccharides are the main effective components of Cynomorium songaricum's stem that perform biological activities and have positive impacts on immune enhancement. In this study, the polysaccharide CSP-III of Cynomorium songaricum's stem was isolated using a DEAE-52 cellulose column through Sephadex G-100 gel column chromatography. Upon analysis, the monosaccharide composition of CSP-III included Mannose (Man), Glucuronic acid (GlcA), Galacturonic acid (GalA), Rhamnose (Rha), Glucose (Glc), Galactose (Gal), and Arabinose (Ara), at a molar ratio of 0.01:0.11:0.03:0.57:0.02:0.32:1. The molecular weight of CSP-III was 4018234 Da. Meanwhile, the capacity of CSP-III, at various concentrations, to stimulate the proliferation of mouse spleen lymphocytes in vitro was compared, and the influence of CSP-III on cell proliferation was examined using RAW264.7 mouse mononuclear macrophages as a model. The influence of CSP-III on the expression of important phosphorylating proteins in the MAPK signaling pathway was initially analyzed by Western blotting. In RAW264.7 cells, CSP-III promoted the phosphorylation of JNK proteins, which thus activated the MAPK signaling cascade and exerted immunomodulatory effects. Moreover, according to in vivo studies using cyclophosphamide (CTX)-induced immunosuppression mouse models, CSP-III improved the CTX-induced histopathological damage, promoted T and B lymphocyte proliferation, upregulated CD4+ and CD8+ T-lymphocyte counts in the spleen, increased the serum levels of IgG and IgM, and activated three essential proteins of the MAPK signaling pathway. As revealed by analysis of intestinal flora, CSP-III improved the immune function by maintaining the homeostasis of the bacterial flora by boosting the relative abundances of some beneficial bacterial groups, such as Bacteroidetes, Desmodium, and Actinomyces, and reducing the relative abundance of Aspergillus phylum. Through in vitro and in vivo experiments, our present study demonstrates that polysaccharides from the stem of Cynomorium songaricum possess strong immunoregulatory effects. Findings in this work provide theoretical support for the potential application of Cynomorium songaricum in the field of health food.

Immunization of BALB/c Mice with Killed Tachyzoites of Toxoplasma gondii against Acute Toxoplasmosis.

Toxoplasma gondii with widespread distribution infects over one third of human populations in the world and can cause serious life-threatening diseases especially for the immunodeficient patients in acute toxoplasmosis. As the clinical pharmaceutical drugs with severe side effects for treatment and non-ideal extant vaccines for prevention, more work starves to be done for keeping advantages in the athletics. Aluminum adjuvant and hybrid formaldehyde-killed tachyzoites of T. gondii RH and GT1 isolates were prepared to intramuscularly immunize BALB/c mice for five times at 0, 3, 7, 14 and 21 days post first injection. The triggered humoral and cellular immune responses at two weeks post the last immunization and the survival times of infected mice were examined for the hybrid immunization scheme judgement. The anti-RH and anti-GT1 specific antibodies were both increased at one week prior to challenge (P < 0.05), and the survival times of immunized mice (7.33 ± 0.71 d for RH, 7.22 ± 0.97 d for GT1) against acute toxoplasmosis were significantly prolonged by the immunizations performed in the study compared to blank control (6.67 ± 0.50 d for RH, 6.33 ± 0.71 d for GT1; P < 0.05), with the higher IFN-γ, IL-2 and IL-12p70 in sera, the elevated CD3e+CD4+ T and CD3e+CD8a+ T cells, and the enhanced lymphocyte proliferation in spleen (P < 0.05). The hybrid killed tachyzoites with aluminum adjuvant induced humoral and cellular immune responses of mice, and offered mildly protective efficacy against acute toxoplasmosis.

Intrapulmonary IFN-γ instillation causes chronic lymphocytic inflammation in the spleen and lung through the CXCR3 pathway

Some patients with chronic refractory cough have high levels of pulmonary IFN-γ and IFN-γ-producing T lymphocytes. Pulmonary IFN-γ administration causes acute airway lymphocytic inflammation and cough hypersensitivity by increasing the number of pulmonary IFN-γ-producing T lymphocytes, but these lymphocytes may be recruited from other organs. Intraperitoneal IFN-γ injection can increase the spleen weight of mice. It remains elusive whether pulmonary IFN-γ can induce chronic airway lymphocytic inflammation and cough hypersensitivity by stimulating the proliferation of IFN-γ -producing T lymphocytes in the spleen. Here, we found that pulmonary IFN-γ administration induced chronic airway inflammation and chronic cough hypersensitivity with an increased number of IFN-γ-producing T lymphocytes in the spleen, blood and lung. Pulmonary IFN-γ administration also increased 1) the proliferation of spleen lymphocytes in vivo and 2) the IP-10 level and CXCR3+ T lymphocyte numbers in the spleen and lung of mice. IP-10 could promote the proliferation of spleen lymphocytes in vitro but not blood lymphocytes or lung-resident lymphocytes. AMG487, a potent inhibitor of binding between IP-10 and CXCR3, could block pulmonary IFN-γ instillation-induced chronic airway lymphocytic inflammation and the proliferation of IFN-γ-producing T lymphocytes in mouse spleens. In conclusion, intrapulmonary IFN-γ instillation may induce the proliferation of splenic IFN-γ-producing T lymphocytes through IP-10 and the CXCR3 pathway. The IFN-γ-producing T lymphocytes in blood, partly released from the mouse spleen, may be partly attracted to the lung by pulmonary IP-10 through the CXCR3 pathway. IFN-γ-producing T lymphocytes and IFN-γ in the lung may cause chronic airway lymphocytic inflammation and chronic cough hypersensitivity.

Nanoparticulate impurities in the pharmaceutical excipient trehalose induce an early immune response

BackgroundPharmaceutical excipients are an important part of biological products. However, few attempts have been made to distinguish between the risk of inflammation associated with the biological products themselves and that associated with excipients. The analysis of early immune response risk associated with excipients added to biological products is an important step in exploring the complex mechanism of side effects in susceptible patients. Methods and ResultsIn this study, nanoparticle impurities (NPIs) were extracted from trehalose and characterized. A mouse popliteal lymph node cell (PLNA) model, a mouse spleen lymphocyte model, a human peripheral blood mononuclear cell cytokine release model, and a macrophage complement activation model were established to comprehensively evaluate the early immune risk related to impurities in the trehalose excipient. Although popliteal lymph node cell counts in mice did not show significant differences, all other models indicated possible immune risk. In the PLNA model, NPIs caused significant toe thickening in mice, whereby the content of IgE and MCP-1 increased significantly. NPIs significantly increased the proliferation and differentiation of spleen lymphocytes according to the CCK-8 assay and flow cytometry. After treatment with NPIs, the release of IgE and a variety of cytokines (MIP-1α, IFN-γ, IL-2, IL-8, TNF-α, IL-6, IL-1α) in human peripheral blood cells was significantly increased according to ELISA, while a concomitant increase of C3a/C5a as well as C4a/Bb proved that NPIs activated the complement system. ConclusionNPIs from trehalose elicited an immune response in vitro, and the immune response to trehalose may be related to NPIs and not the excipient itself. Different batches of trehalose showed different immune response effects. The currents research suggests that when trehalose is applied in high-risk administration routes, NPIs should be assessed and reasonably controlled.

Evaluation of the Quality of Codonopsis Radix in Different Growth Years by the AHP-CRITIC Method.

The quality of traditional Chinese medicines (TCM) has a significant correlation with the source and growth years. However, there is no research on the relationship between the growth period and the quality of Codonopsis Radix (CR). This work aims to evaluate the quality of CR in different growth years (2-5 years). First, the content of 6 efficacy-related and 28 nutrient-related components in different growth years of CR was analyzed. The results showed that with the increase in growth years of CR, the content of some components increased, while some decreased. Then, the AHP-CRITIC method was performed to score the CR in different growth years, the results showed that the comprehensive score of CR increased with the increase of growth years, and the 5-year-CR had the highest score. Finally, in vitro activity assays were designed to verify the evaluation results. The results demonstrated that with the increase of growth years, the spleen lymphocyte proliferation activity and DPPH free radical scavenging activity of CR were enhanced, which proved that the AHP-CRITIC method is reasonable to evaluate the quality of CR in different growth years. The aforementioned findings demonstrated that CR quality improved with longer planting years.

Staphylococcal enterotoxin B as DNA vaccine against breast cancer in a murine model.

Recently, many efforts have been made to treat cancer using recombinant bacterial toxins and this strategy has been used in clinical trials of various cancers. Therapeutic DNA cancer vaccines are now considered as a promising strategy to activate the immune system against cancer. Cancer vaccines could induce specific and long-lasting immune responses against tumors. This study aimed to evaluate the antitumor potency of the SEB DNA vaccine as a new antitumor candidate against breast tumors in vivo. To determine the effect of the SEB construct on inhibiting tumor cell growth in vivo, the synthetic SEB gene, subsequent codon optimization, and embedding the cleavage sites were sub-cloned to an expression vector. Then, SEB construct, SEB, and PBS were injected into the mice. After being vaccinated, 4T1 cancer cells were injected subcutaneously into the right flank of mice. Then, the cytokine levels of IL-4 and IFN-γ were estimated by the ELISA method to evaluate the antitumor activity. The spleen lymphocyte proliferation, tumor size, and survival time were assessed. The concentration of IFN-γ in the SEB-Vac group showed a significant increase compared to other groups. The production of IL-4 in the group that received the DNA vaccine did not change significantly compared to the control group. The lymphocyte proliferation increased significantly in the mice group that received SEB construct than PBS control group (p < 0.001). While there was a meaningful decrease in tumor size (p < 0.001), a significant increase in tumor tissue necrosis (p < 0.01) and also in survival time of the animal model receiving the recombinant construct was observed. The designed SEB gene construct can be a new model vaccine for breast cancer because it effectively induces necrosis and produces specific immune responses. This structure does not hurt normal cells and is a safer treatment than chemotherapy and radiation therapy. Its slow and long-term release gently stimulates the immune system and cellular memory. It could be applied as a new model for inducing apoptosis and antitumor immunity to treat cancer.

Study on the preparation and biological activities of low molecular weight squid ink polysaccharide from Sepiella maindroni

Sepiella maindroni ink polysaccharide (SIP) from the ink of cuttlefish Sepiella maindroni and its sulfated derivative (SIP-SII) have been demonstrated to possess diverse biological activities. But little is known about low molecular weight squid ink polysaccharides (LMWSIPs). In this study, LMWSIPs were prepared by acidolysis, and the fragments with molecular weight (Mw) distribution in the ranges of 7 kDa to 9 kDa, 5 kDa to 7 kDa and 3 kDa to 5 kDa were grouped and named as LMWSIP-1, LMWSIP-2 and LMWSIP-3, respectively. The structural features of LMWSIPs were elucidated, and their anti-tumor, antioxidant and immunomodulatory activities were also studied. The results showed that with the exception of LMWSIP-3, the main structures of LMWSIP-1 and LMWSIP-2 did not change compared with SIP. Though there were no significant differences in the antioxidant capacity between LMWSIPs and SIP, the anti-tumor and immunomodulatory activities of SIP were enhanced to a certain extent after degradation. It is particularly noteworthy that the activities of LMWSIP-2 in anti-proliferation, promoting apoptosis and inhibiting migration of tumor cells as well as promoting the proliferation of spleen lymphocytes were significantly higher than those of SIP and the other degradation products, which is promising in the anti-tumor pharmaceutical field.

Selenium Nanoparticles Based on Morinda officinalis Polysaccharides: Characterization, Anti-Cancer Activities, and Immune-Enhancing Activities Evaluation In Vitro

Recently, selenium nanoparticles have been drawing attention worldwide, and it is crucial to increase the stability of nano-Se. Morinda officinalis polysaccharides (MOP) are the main active component in Morinda officinalis radix. However, their low activity has limited their application. A novel selenium nanoparticle (Se-MOP) was prepared to solve these problems using MOP as a dispersant. The zeta potential was measured to evaluate the stability, and UV and ATR-FTIR were used to investigate the binding type of selenium and MOP. The morphology was observed by the TEM method. Furthermore, the inhibitory effect on five selected cancer cells (HepG2, MCF-7, AGS, PC9, and HCT8) was evaluated, showing remarkable inhibition of all five cancer cells. The mechanism of inhibition was also investigated by cell circle assay, and it was found that Se-MOP could induce cell circle G0/G1 phase arrest. Immune-enhancing activities were evaluated by measuring the proliferation and cytokines of mouse spleen lymphocytes in vitro and quantitative RT-PCR. The results indicated that single stimulation of Se-MOP and synergistic stimulation with PHA or LPS increased immune capacity and improved immune by increasing the expression of cytokines.

Immunomodulatory effect of ethanol-soluble oligopeptides from Atlantic cod (Gadus morhua)

There are many active substances in Atlantic cod (Gadus morhua) explaining the variety of biological activities. In order to study the immunomodulatory activity and the mechanism of Atlantic cod peptides at the cellular level. In this study, cod peptides were isolated by 80 % ethanol extraction method, the isolated ethanol-soluble cod peptides (CP-ES) were investigated and their immunomodulatory activity was verified. Additionally, CP-ES showed lower molecular weight and more hydrophobic amino acids. CP-ES could promote the proliferation of spleen lymphocytes and T lymphocytes in mice, suggesting that CP-ES may regulate adaptive immunity. It promoted the release of NO and the expression of iNOS, TNF-α, IL-6 and IL-1β genes in macrophages, suggesting that CP-ES may regulate innate immunity. CP-ES could promote the expression of TLR2 gene, and the peptides identified in CP-ES were docked with TLR2 to predict the peptides playing a major role in CP-ES. These results suggested that CP-ES may regulate the immune activity of both innate and adaptive lines.

Early-life maternal deprivation affects the mother-offspring relationship in domestic pigs, as well as the neuroendocrine development and coping behavior of piglets.

Early-life adversity may have programming effects on the psychological and physiological development of offspring. Domestic pigs (Sus scrofa) are an excellent model species for studying these effects because of their many physiological similarities to humans. Piglets from 10 sows were subjected to daily 2-h maternal deprivation on postnatal days (PND) 2–15 alone (DA) or in a group of littermates (DG). Control piglets (C) from 10 sows stayed with their mothers. Mother-offspring interaction, milk oxytocin, and cortisol were analyzed. An open-field/novel-object (OF/NO) test was performed with piglets on PNDs 16 and 40. Plasma cortisol and immune parameters were determined on PND 5 and 16. Two piglets from each group and sow were sacrificed on PND 20 and stress-related gene expression in the limbic system and prefrontal cortex (PFC), as well as splenic lymphocyte proliferative abilities, were examined. The milk cortisol of sows increased during the first separation of mother and offspring on the second day of lactation, whereas milk oxytocin did not change. The increase in cortisol by the OF/NO test on PND 16 was greater in C piglets than in DA and DG ones. DA piglets showed less agitated behavior than DG and C piglets in the OF/NO test at PND 16, but appeared more fearful. On PND 40, DA piglets showed more arousal than DG and C piglets in the OF/NO test. Neither plasma IgA nor N/L ratios in blood nor mitogen-induced proliferation of spleen lymphocytes were affected by deprivation. We found a higher mRNA expression of CRHR1 in the hypothalamus and a higher expression of MR in the hippocampus in DA piglets than in DG ones. The expression of GR, MR, and CRHR1 genes in the PFC was reduced by maternal deprivation, however, the expression of arginine vasopressin and oxytocin receptors was not affected. Repeated maternal deprivation induces sustained effects on stress reactivity and behavior of domestic piglets. Some of these effects were buffered by the presence of littermates. In addition, we found sex-specific differences in behavior and gene expression.

Tim3scFv-Transforming Lactobacillus Inhibits Transplanted Tumor of Renal-Cell Carcinoma

Introduction: T-cell immunoglobulin-3 (Tim-3) antibody drugs can treat malignant renal tumors but are expensive. To overcome this limitation, Lactococcus lactis host bacteria were used to express Tim-3 single-chain antibodies. Methods: The pLAN-CTB-Tim3scFv plasmid was constructed using molecular cloning technology and transformed into Lactococcus lactis. Expression and immune activity of proteins in the transformed bacteria were analyzed using Western blotting and enzyme-linked immunosorbent assay in vitro. A mouse subcutaneously transplanted tumor model of renal adenocarcinoma was constructed. The promoting effect of transformed bacteria on mouse spleen lymphocyte activation and their inhibitory effect on transplanted tumors were analyzed. Results: Transformed L. lactis NZ-CTB-Tim3scFv and NZ-Tim3scFv were successfully constructed. CTB-Tim3scFv secreted by NZ-CTB-Tim3scFv showed immunological activity. Compared with the NZ-Tim3scFv and NZ-Vector groups, the subgroups of splenic lymphocytes in the NZ-CTB-Tim3scFv group had a higher proportion of CD3⁺CD4⁺, CD3⁺CD8a⁺, and CD3⁺CD69⁺ cells. Ki67 and CD31 expression in the NZ-CTB-Tim3scFv group was significantly reduced. Tumor volume in the NZ-CTB-Tim3scFv group increased the least. Discussion/Conclusion: Secretion of CTB-Tim3scFv promoted the proliferation and activation of spleen lymphocytes and inhibited growth, cell proliferation, and angiogenesis of tumors. The proposed method is low cost and convenient with potential to become a new immunotherapy approach for renal-cell carcinoma.

Immunosuppressive Cytochalasins from the Mangrove Endophytic Fungus Phomopsis asparagi DHS-48.

Three new cytochalasins, phomoparagins A-C (1–3), along with five known analogs (4–8), were isolated from Phomopsis asparagi DHS-48, a mangrove-derived endophytic fungus. Their structures, including their absolute configurations, were elucidated using a combination of detailed HRESIMS, NMR, and ECD techniques. Notably, 1 possessed an unprecedented 5/6/5/8/5-fused pentacyclic skeleton. These compounds were tested for their inhibitory activity against concanavalin A (ConA)/lipopolysaccharide (LPS)-induced spleen lymphocyte proliferation and calcineurin (CN) enzyme. Several metabolites (2 and 4–6) exhibited fascinating inhibitory activities with a relatively low toxicity. Furthermore, 2 was demonstrated to inhibit ConA-stimulated activation of NFAT1 dephosphorylation and block NFAT1 translocation in vitro, subsequently inhibiting the transcription of interleukin-2 (IL-2). Our results provide evidence that 2 may, at least partially, suppress the activation of spleen lymphocytes via the CN/NFAT signaling pathway, highlighting that it could serve as an effective immunosuppressant that is noncytotoxic and natural.

Interferon-γ enhances the immunosuppressive ability of canine bone marrow-derived mesenchymal stem cells by activating the TLR3-dependent IDO/kynurenine pathway.

The immunomodulatory function of mesenchymal stem cells (MSCs) has been considered to be vital for MSC-based therapies. Many works have been devoted to excavate effective strategies for enhancing the immunomodulation effect of MSCs. Nonetheless, canine MSC-mediated immunomodulation is still poorly understood. The inflammatory microenvironment was simulated through the employment of interferon-γ (IFN-γ) in a culture system. Compared with unstimulated cBMSCs, IFN-γ stimulation increased the mRNA levels of Toll-like receptor 3 (TLR3) and indoleamine 2, 3-dioxygenase 1 (IDO-1), and simultaneously enhanced the secretion of immunosuppressive molecules, including interleukin (IL)-10, hepatocyte growth factor (HGF), and kynurenine in cBMSCs. IFN-γ stimulation significantly enhanced the ability of cBMSCs and their supernatant to suppress the proliferation of murine spleen lymphocytes. Lymphocyte subtyping evaluation revealed that cBMSCs and their supernatant diminished the percentage of CD3+CD4+ and CD3+CD8+ lymphocytes compared with the control group, with a decreasing CD4+/CD8+ ratio. Notably, exposure to IFN-γ decreased the CD4+/CD8+ ratio more effectively than unstimulated cells or supernatant. Additionally, IFN-γ-stimulation increased the mRNA levels of the Th1 cytokines TNF-α, and remarkably decreased the mRNA level of the Th2 cytokine IL-4 and IL-10. Our findings substantiate that IFN-γ stimulation can enhance the immunomodulatory properties of cBMSCs by promoting TLR3-dependent activation of the IDO/kynurenine pathway, increasing the secretion of immunoregulatory molecules and strengthening interactions with T lymphocytes, which may provide a meaningful strategy for the clinical application of cBMSCs in immune-related diseases.