Zhen-Wu-Tang ameliorates lupus nephritis by diminishing renal tissue-resident memory CD8+ T cells via suppressing IL-15/STAT3 pathway.

Zhen-Wu-Tang (ZWT), a conventional herbal mixture, has been recommended for treating lupus nephritis (LN) in clinic. However, its mechanisms of action remain unknown. Here we aimed to define the immunological mechanisms underlying the effects of ZWT on LN and to determine whether it affects renal tissue-resident memory T (TRM) cells. Murine LN was induced by a single injection of pristane, while in vitro TRM cells differentiated with IL-15/TGF-ß. We found that ZWT or mycophenolate mofetil treatment significantly ameliorated kidney injury in LN mice by decreasing 24-h urine protein, Scr and anti-dsDNA Ab. ZWT also improved renal pathology and decreased IgG and C3 depositions. In addition, ZWT down-regulated renal Desmin expression. Moreover, it lowered the numbers of CD8+ TRM cells in kidney of mice with LN while decreasing their expression of TNF-α and IFN-γ. Consistent with in vivo results, ZWT-containing serum inhibited TRM cell differentiation induced by IL-15/TGF-ß in vitro. Mechanistically, it suppressed phosphorylation of STAT3 and CD122 （IL2/IL-15Rß）expression in CD8+ TRM cells. Importantly, ZWT reduced the number of total F4/80+CD11b+ and CD86+, but not CD206+, macrophages in the kidney of LN mice. Interestingly, ZWT suppressed IL-15 protein expression in macrophages in vivo and in vitro. Thus, we have provided the first evidence that ZWT decoction can be used to improve the outcome of LN by reducing CD8+ TRM cells via inhibition of IL-15/IL-15R /STAT3 signaling.

Methotrexate and electrostimulation cooperate to alleviate the relapse of psoriasiform skin inflammation by suppressing memory T cells.

Methotrexate (MTX) is an immunosuppressant used to treat autoimmune diseases, including psoriasis. However, like other immunosuppressants, MTX alone does not prevent their recurrence. Electrostimulation (ES) has been utilized to treat some inflammatory disorders without any major side-effect. But it remains unknown if ES alone, or together with MTX, ameliorates autoimmune disease relapse: a sticky medical problem. In particular, the mechanisms underlying ES action remain unclear. The objective of this study was to determine an impact of ES and/or MTX on psoriasis relapse and their potential cooperation. We found that regional ES, but not MTX, ameliorated psoriasiform skin inflammation recurrence. Interestingly, treatment with both MTX and ES further prevented psoriasis recurrence compared to ES alone. Moreover, ES downregulated potassium channel Kv1.3 on T-cells and reduced CD4+/CD8+ effector memory (TEM) and CD8+ skin-resident memory T (TRM) cells, while ES plus MTX further decreased CD8+ TEM/TRM cells compared to ES alone. However, ES failed to further attenuate psoriasis recurrence or suppress T cell memory in Kv1.3-deficient mice, whereas lack of Kv1.3 itself ameliorated psoriasis relapse by shrinking T cell memory pool. Importantly, ES moderately inhibited T-cell proliferation in vitro. ES also reduced human CD8+ TRM cells and attenuated human skin lesions in humanized mice grafted with lesional skin from patients with recurrent psoriasis, with an enhanced efficacy in mice treated with both ES and MTX. Thus, ES and MTX cooperated to prevent psoriasis relapse by reducing T-cell memory via targeting potassium channel Kv1.3. Our studies may be implicated for treating human psoriasis.

Hyperoside Suppresses Renal Inflammation by Regulating Macrophage Polarization in Mice With Type 2 Diabetes Mellitus.

Accumulating evidence reveals that both inflammation and lymphocyte dysfunction play a vital role in the development of diabetic nephropathy (DN). Hyperoside (HPS) or quercetin-3-O-galactoside is an active flavonoid glycoside mainly found in the Chinese herbal medicine Tu-Si-Zi. Although HPS has a variety of pharmacological effects, including anti-oxidative and anti-apoptotic activities as well as podocyte-protective effects, its underlying anti-inflammatory mechanisms remain unclear. Herein, we investigated the therapeutic effects of HPS on murine DN and the potential mechanisms responsible for its efficacy. We used C57BLKS/6J Lepdb/db mice and a high glucose (HG)-induced bone marrow-derived macrophage (BMDM) polarization system to investigate the potentially protective effects of HPS on DN. Our results showed that HPS markedly reduced diabetes-induced albuminuria and glomerular mesangial matrix expansion, accompanied with a significant improvement of fasting blood glucose level, hyperlipidaemia and body weight. Mechanistically, pretreatment with HPS effectively regulated macrophage polarization by shifting proinflammatory M1 macrophages (F4/80+CD11b+CD86+) to anti-inflammatory M2 ones (F4/80+CD11b+CD206+) in vivo and in bone marrow-derived macrophages (BMDMs) in vitro, resulting in the inhibition of renal proinflammatory macrophage infiltration and the reduction in expression of monocyte chemoattractant protein-1 (MCP-1), tumor necrosis factor (TNF-α) and inducible nitric oxide synthase (iNOS) while increasing expression of anti-inflammatory cytokine Arg-1 and CD163/CD206 surface molecules. Unexpectedly, pretreatment with HPS suppressed CD4+ T cell proliferation in a coculture model of IL-4-induced M2 macrophages and splenic CD4+ T cells while promoting their differentiation into CD4+IL-4+ Th2 and CD4+Foxp3+ Treg cells. Taken together, we demonstrate that HPS ameliorates murine DN via promoting macrophage polarization from an M1 to M2 phenotype and CD4+ T cell differentiation into Th2 and Treg populations. Our findings may be implicated for the treatment of DN in clinic.

Dihydroartemisinin ameliorates psoriatic skin inflammation and its relapse by diminishing CD8+ T-cell memory in wild-type and humanized mice.

Conventional immunosuppressants cause side effects and do not prevent the recurrence of autoimmune diseases. Moreover, they may not inhibit autoimmunity mediated by pathogenic memory T-cells. Dihydroartemisinin (DHA) has been shown to regulate autoimmunity. However, it remains unknown whether DHA impacts psoriasis and its recurrence. The objective of this study was to determine therapeutic effects of DHA on psoriasis and its relapse as well as its underlying mechanisms. Methods: We established animal models of imiquimod (IMQ)-induced psoriasis-like wild-type mice and humanized NSG mice receiving lesional human skin from patients with psoriasis. Many immunoassays, including immunohistochemistry, flow cytometry, quantitative RT-PCR and Western blotting, were performed. Results: We found that DHA not only ameliorated acute skin lesion of psoriatic mice, but also alleviated its recurrence by diminishing CD8+ central memory T (TCM) and CD8+ resident memory T (TRM) cells. It attenuated epidermal pathology and T-cell infiltration in the skin of IMQ-induced psoriatic mice while suppressing expression of IL-15, IL-17 and other proinflammatory cytokines in the skin. Surprisingly, DHA reduced the frequency and number of CD8+, but not CD4+, subset of CD44highCD62Lhigh TCM in psoriatic mice, whereas methotrexate (MTX) lowered CD4+, but not CD8+, TCM frequency and number. Indeed, DHA, but not MTX, downregulated eomesodermin (EOMES) and BCL-6 expression in CD8+ T-cells. Furthermore, DHA, but not MTX, reduced the presence of CD8+CLA+, CD8+CD69+ or CD8+CD103+ TRM cells in mouse skin. Interestingly, treatment with DHA, but not MTX, during the first onset of psoriasis largely prevented psoriasis relapse induced by low doses of IMQ two weeks later. Administration of recombinant IL-15 or CD8+, but not CD4+, TCM cells resulted in complete recurrence of psoriasis in mice previously treated with DHA. Finally, we demonstrated that DHA alleviated psoriatic human skin lesions in humanized NSG mice grafted with lesional skin from psoriatic patients while reducing human CD8+ TCM and CD103+ TRM cells in humanized mice. Conclusion: We have provided the first evidence that DHA is advantageous over MTX in preventing psoriasis relapse by reducing memory CD8+ T-cells.

Antitumor effects of immunity-enhancing traditional Chinese medicine.

Traditional Chinese Medicine (TCM) has been traditionally used to treat patients with cancers in China. It not only alleviates the symptoms of tumor patients and improves their quality of life, but also controls the size of tumors and prolongs the survival of tumor patients. While some herbs of TCM may exert therapeutic effects by directly targeting cancer cells or reducing side effects caused by antitumor drugs, others can control tumor growth and metastasis via enhancing antitumor immunity. In particular, TCM can exert antitumor effects by upregulating immune responses even in immunosuppressive tumor microenvironment. For instance, it reduces the number of M2-type macrophages and Treg cells in the tumor tissue. Although extensive reviews on directly killing cancer cells by TCM have been conducted, a review of anticancer activity of TCM solely based on its immunity-enhancing capacity is unusual. This review will summarize research progress of antitumor TCM that regulates the immune system, including both innate immunity, such as macrophages, dendritic cells, natural killer cells and MDSCs, and adaptive immunity, including CD4+/CD8+ T lymphocytes, regulatory T cells (Tregs) and B cells. As cancer immunotherapy has recently achieved certain success, it is expected that the clinical applications of immunity-enhancing TCM or traditional medicine for treating various cancer patients will be expanded. Further studies on the mechanisms by which TCM regulates immunity will provide new insights into how TCM controls tumor growth and metastasis, and may help improve its therapeutic effects on various cancers in clinic.

Cordyceps militaris polysaccharides exerted protective effects on diabetic nephropathy in mice via regulation of autophagy.

The present study was designed to investigate the protective effects of Cordyceps militaris polysaccharides (CMP) on STZ-treated DN mice. CMP were identified by FT-IR and HPLC. Diabetic nephropathy (DN) was induced in male C57BL/6 mice by the injection of streptozotocin (STZ, 50 mg kg-1) in citrate buffer on 5 consecutive days. Administration of CMP at 200 and 400 mg kg-1 or irbesartan at 60 mg kg-1 in the STZ-treated mice could prevent the damage caused by STZ. CMP significantly reduced the STZ-induced higher expression of the kidney index, TC, TG, MDA, urinary protein, Scr, and BUN, while it markedly increased the STZ-induced decrease in GSH levels compared with the DN group. Histopathology analysis of the kidney by PAS, Masson, and HE staining confirmed the renal injury induced by STZ and the protective effects of CMP. Transmission electron microscopy (TEM) results confirmed the severe foot process effacement induced by STZ, but CMP treatment inhibited the podocytes' structure defects and ameliorated the function of podocytes. Desmin was measured by immunofluorescence and was related to podocyte injury. The results showed that CMP lessened the expression of desmin induced by STZ. CD68 expression was measured by immunohistochemistry analysis, and the expressions of IL-1ß, IL-6, and MCP-1 mRNA were measured by qRT-PCR. The results showed that CMP suppressed the expressions of CD68, IL-1ß, IL-6, and MCP-1 mRNA induced by STZ. The role of autophagy in the treatment of DN mice with CMP was detected by TEM and western blotting. The results showed that the administration of CMP was able to overcome the STZ-treated autophagy deficiency, significantly increase the rate of autophagy in the kidney, promote the expression of Atg5, beclin1 and LC3 protein, and reduce the expression of p62 protein. In conclusion, the present study demonstrates that CMP exert a protective effect on DN in STZ-treated mice possibly via activation of autophagy.

Renoprotective effects of artemisinin and hydroxychloroquine combination therapy on IgA nephropathy via suppressing NF-κB signaling and NLRP3 inflammasome activation by exosomes in rats.

Immunoglobulin A nephropathy (IgAN) is an autoimmune kidney disease with complex pathogenesis leading to end-stage renal damage. The prime pathological characteristics of IgAN are IgA immune complexes deposition accompany with mesangial cell proliferation and urine protein elevation. Artemisinin (ART) is extracted from traditional Chinese medicine Artemisia annua L. Hydroxychloroquine (HCQ) is a classical antimalarial drug applied in the treatment of autoimmune diseases. Both of them possess anti-inflammatory and immunomodulatory properties. The purpose of this research was to investigate the pharmacological effects of ART combined with HCQ (AH) and discuss thoroughly the potential molecular mechanisms in IgAN. In vivo, our results demonstrated that AH could efficiently ameliorate kidney damage by improving kidney dysfunction and reducing the levels of 24 h urine protein, IgA and IgG immune complexes deposition in glomerulus of IgAN rats. Interestingly, AH obviously promoted the secretion of exosomes in renal tissues, inhibited the expressions of nuclear factor-κB (NF-κB) signaling and NLRP3 inflammasome-related proteins, including IκB-α, p-p65, NLRP3, ASC, IL-1ß and caspase-1 in IgAN rats. In vitro, further mechanistic study illustrated that exosomes derived from human renal tubular epithelial cells (HK-2) were significantly enhanced by AH, which could be utterly taken up in human mesangial cells (HMCs) and inhibited the activation of NF-κB pathway and NLRP3 inflammasome after AH intervention. However, GW4869 interdicted the promotive effect of AH on exosomes from HK-2 cells and the suppression of exosomes on NF-κB/NLRP3 activation in HMCs. Taken together, this study demonstrated that there was an inhibitory effect of AH therapy on NF-κB/NLRP3 signaling via mediating exosomes release in IgAN rats, which provided an alternative approach for IgAN treatment.

Zhen-wu-tang ameliorates membranous nephropathy rats through inhibiting NF-κB pathway and NLRP3 inflammasome.

BACKGROUND: Zhen-wu-tang (ZWT), a traditional herbal formula, has been widely used for the treatment of kidney diseases in clinics, but the underlying molecular mechanisms have not been fully understood. PURPOSE: Inflammation mediated podocyte injury has been reported to constitute a crucial part in the pathogenesis of membranous nephropathy (MN). The current study was designed to evaluate the effect of ZWT on MN related to nuclear factor-κB (NF-κB) pathway and NLRP3 inflammasome. METHODS: The main components of ZWT were identified by 3D-ultra performance liquid chromatography (3D-UPLC) assay. A MN rat model induced by cationic-bovine serum albumin (C-BSA) and podocytes stimulated by TNF-α were used in this study. The 24 h urine protein, serum total cholesterol (TC) and triglyceride (TG), as well as kidney histology were measured to evaluate kidney damage. The expressions of IgG and complement 3 (C3), and the co-localization of NLRP3 and ASC were detected by immunofluorescence. The expressions of podocyte injury related protein desmin, podocin were measured by immunohistochemistry and western blot. Cell vitality of cultured podocytes was detected by MTT assay, as apoptosis assay was measured via flow cytometry. The protein expressions of p-p65, p-IκBα, NLRP3, Caspase-1, IL-1ß were detected by western blot. RESULTS: Our results showed that ZWT significantly ameliorated kidney damage in MN model rats by decreasing the levels of 24 h urine protein, TC and TG. ZWT also improved renal histology and reduced the expressions of IgG and C3 in glomerulus. In addition, ZWT lessened the expressions of desmin, but increased podocin expression in vivo and vitro. ZWT protected cultured podocytes by maintaining cell vitality and inhibiting apoptosis. Moreover, we found that ZWT suppressed the expressions of NLRP3, Caspase-1, IL-1ß and the co-localization of NLRP3 and ASC. Furthermore, the inhibition of NLRP3 inflammasome under ZWT treatment were accompanied by down-regulation of NF-κB pathway, as the p-p65 and p-IκBα protein expression were reduced. CONCLUSIONS: Our present study indicates that the inhibition of NF-κB pathway and NLRP3 inflammasome might be the potential mechanisms for the therapeutic effects of ZWT against MN.

Immunosuppressive effect of artemisinin and hydroxychloroquine combination therapy on IgA nephropathy via regulating the differentiation of CD4+ T cell subsets in rats.

Immunoglobulin A nephropathy (IgAN) is an autoimmune kidney disease with complex pathogenesis leading to end-stage renal damage. The crucial pathological characteristic in IgAN is IgA immune complexes deposition accompany with mesangial cell proliferation and mesangial matrix expansion. Artemisinin (ART) is isolated from traditional Chinese medicine Artemisia annua L. Hydroxychloroquine (HCQ) is a classical antimalarial drug used to treat autoimmune diseases. Both of them possess immunosuppressive, immunomodulatory and anti-inflammatory features. The aim of this study was to investigate the pharmacological effects of ART combined with HCQ (AH) and explore the underlying mechanisms in IgAN. In vivo, our results showed that AH could significantly improve kidney dysfunction, decrease mesangial matrix expansion as well as immune complexes in mesangial area visualized by H&E and PAS staining. The depositions of IgA immune complexes and complement 3 (C3) were obviously reduced after AH treatment by immunofluorescence. Interestingly, the morphology of kidney and spleen was significantly swelled but reverted by AH in IgAN rats. Further mechanistic study showed that the higher proportions of the Th2 and Th17 cells were reduced but the lower differentiation of Th1 and Treg cells subsets were promoted by AH. Taken together, this study demonstrated that there was an immunosuppressive effect of AH therapy on IgAN rats via regulating the differentiation of CD4+ T cell subsets, which provided an alternative approach for IgAN treatment.

Bacterial Community Assembly in a Typical Estuarine Marsh with Multiple Environmental Gradients.

Bacterial communities play essential roles in estuarine marsh ecosystems, but the interplay of ecological processes underlying their community assembly is poorly understood. Here, we studied the sediment bacterial communities along a linear gradient extending from the water-land junction toward a high marsh, using 16S rRNA gene amplicon sequencing. Bacterial community compositions differed significantly between sediment transects. Physicochemical properties, particularly sediment nutrient levels (i.e., total nitrogen [TN] and available phosphorus [AP]), as well as sediment physical structure and pH (P < 0.05), were strongly associated with the overall community variations. In addition, the topological properties of bacterial cooccurrence networks varied with distance to the water-land junction. Both node- and network-level topological features revealed that the bacterial network of sediments farthest from the junction was less intense in complexity and interactions than other sediments. Phylogenetic null modeling analysis showed a progressive transition from stochastic to deterministic community assembly for the water-land junction sites toward the emerging terrestrial system. Taken together, data from this study provide a detailed outline of the distribution pattern of the sediment bacterial community across an estuarine marsh and inform the mechanisms and processes mediating bacterial community assembly in marsh soils.IMPORTANCE Salt marshes represent highly dynamic ecosystems where the atmosphere, continents, and the ocean interact. The bacterial distribution in this ecosystem is of great ecological concern, as it provides essential functions acting on ecosystem services. However, ecological processes mediating bacterial assembly are poorly understood for salt marshes, especially the ones located in estuaries. In this study, the distribution and assembly of bacterial communities in an estuarine marsh located in south Hangzhou Bay were investigated. The results revealed an intricate interplay between stochastic and deterministic processes mediating the assembly of bacterial communities in the studied gradient system. Collectively, our findings illustrate the main drivers of community assembly, taking into consideration changes in sediment abiotic variables and potential biotic interactions. Thus, we offer new insights into estuarine bacterial communities and illustrate the interplay of ecological processes shaping the assembly of bacterial communities in estuarine marsh ecosystems.

Zhen-wu-tang attenuates Adriamycin-induced nephropathy via regulating AQP2 and miR-92b.

Nephrotic syndrome (NS) is characterized by proteinuria, hypoalbuminemia and edema. The disorder of sodium and water metabolism is a critical mechanism regulating the origination and progression of NS. Zhen-wu-tang (ZWT) has been traditionally used to treat edema disease in China and Japan. The present study was carried out to assess the protective effect of ZWT in Adriamycin-induced (ADR) NS rats and investigate the potential anti-NS mechanisms of ZWT. We found that ZWT treatment ameliorate impaired kidney function and regulate water balance of kidney. Importantly, ZWT increased the expression of Aquaporin-2 (AQP2) which play key roles in maintaining body water homeostasis. Additionally, we determined miRNAs expression patterns in NS rats. Using bioinformatics prediction and miR-92b mimic or inhibitor in vitro, we identified miR-92b as a possible modulator of AQP2. Also we found that ZWT can decrease the expression of miR-92b and reverse the effect of miR-92b on AQP2 in vitro. We further demonstrated that miR-92b directly regulated AQP2 expression by targeting 3'-UTR of AQP2. These finding suggest that ZWT may reduce renal edema in Adriamycin-induced nephropathy via regulating AQP2 and miR-92b.

A Randomized Double-Blind Placebo-Controlled Trial to Evaluate Prophylactic Effect of Traditional Chinese Medicine Supplementing Qi and Hemostasis Formula on Gastrointestinal Bleeding after Percutaneous Coronary Intervention in Patients at High Risks.

OBJECTIVE: To evaluate the clinical efficacy of traditional Chinese medicine (TCM) supplementing Qi and hemostasis formula on gastrointestinal (GI) bleeding after percutaneous coronary intervention (PCI) and thus find out the optimal therapeutic regimen to reduce incidence of GI bleeding without increase of major adverse cardiovascular events (MACEs). METHODS: In the randomized, double-blinded, controlled trial, 117 participants who underwent PCI were enrolled and evenly distributed into treatment arm (59) and control arm (58). Numerous end points were assessed including the primary endpoint of GI bleeding and MACEs and secondary endpoint of thromboelastogram (TEG) (mainly MAadp, inhibition of ADP, and inhibition of AA) and TCM syndrome score during the follow-up phase of 90 days. RESULTS: Incidence of bleeding including GI bleeding and MACE did not differ significantly between two arms (28.82% in treatment arm versus 24.44% in control). However, on both days 30 and 90, TCM treatment remarkably reduced the TCM syndrome total score with notable alteration (P<0.05) except for some parameters such as pulse manifestation. When it came to TEG, however, MAADP increased significantly on day 30 in control arm, accompanied by a notable descending in inhibition rate of ADP pathway (both P<0.01). CONCLUSION: (1) Supplementing Qi and hemostasis formula is equal to Pantoprazole Sodium Enteric-Coated Capsule in hemostasis and gastric mucosal protection; (2) supplementing Qi and hemostasis formula is superior to Pantoprazole Sodium Enteric-Coated Capsule in improving TCM syndrome manifestation possibly through the multitarget mechanism; (3) interference on clopidogrel of supplementing Qi and hemostasis formula might be much less than Pantoprazole Sodium Enteric-Coated Capsule due to the potential CYP450-independent mechanism. This trial is registered with ChiCTR1800014485.

Herbal Components of a Novel Formula PSORI-CM02 Interdependently Suppress Allograft Rejection and Induce CD8+CD122+PD-1+ Regulatory T Cells.

A recipient usually rejects a transplanted organ and thus needs immunosuppressive treatments to prevent rejection. Achieving long-term allograft survival without continuous global immunosuppression is highly desirable in transplantation as long-term immunosuppression causes various side effects. Therefore, it is necessary to search for medicine with potentially less side effects. Traditional Chinese medicine PSORI-CM01 (Yin Xie Ling), a formula with seven natural herbs, has been used to treat patients with psoriasis. Here, we investigated a "sharpened" formula, PSORI-CM02 consisting of only five herbs from PSORI-CM01: Curcumae rhizoma, Radix paeoniae rubra, Rhizoma smilacis glabrae, Mume fructus, and Sarcandrae herba. We examined whether or not PSORI-CM02 would suppress alloimmunity and found that PSORI-CM02 significantly inhibited murine skin allograft rejection and reduced graft-infiltration of CD3+ T cells. Interestingly, omitting any single herbal component rendered the whole formula ineffective in suppression, indicating that these herbal components exert their effects cooperatively as a whole. Moreover, PSORI-CM02 increased CD8+CD122+PD-1+ Treg frequency with CD4+FoxP3+ Tregs remaining unchanged in recipient mice, whereas CsA reduced CD4+FoxP3+ Treg frequency. PSORI-CM02 also hindered CD11c+ DC maturation posttransplantation. Importantly, PSORI-CM02-induced CD8+CD122+PD-1+ Tregs were more potent in suppression of allograft rejection in Rag-/- mice than control Tregs. On the other hand, PSORI-CM02 suppressed T cell proliferation in vitro and reduced their phosphorylation of P70S6K and P50/P65, suggesting that it inhibits both mTOR and NFκB signaling pathways. It also increased IL-10 production while reducing IFNγ level in the supernatant of activated T cells co-cultured with CD8+CD122+PD-1+ Tregs. Furthermore, HPLC fingerprinting ruled out that PSORI-CM02 contained CsA or rapamycin. PSORI-CM02 also did not cause any illness and toxic injury in recipient mice. Thus, we demonstrate that PSORI-CM02 formula suppresses allograft rejection without toxicity.

Chinese medicine Ginseng and Astragalus granules ameliorate autoimmune diabetes by upregulating both CD4+FoxP3+ and CD8+CD122+PD1+ regulatory T cells.

Type 1 diabetes mellitus (T1DM) is an autoimmune disease mainly mediated by effector T cells that are activated by autoantigen, thereby resulting in the destruction of pancreatic islets and deficiency of insulin. Cyclosporine is widely used as an immunosuppressant that suppresses autoimmunity in clinic. However, continuous treatments with conventional immunosuppressive drugs may cause severe side effects. Therefore it is important to seek alternative medicine. Chinese medicine Ginseng and Astragalus granule (GAG) was used to successfully treat type 2 diabetes mellitus in clinic in China. Here we found that GAG ameliorated T1DM in autoimmune NOD mice by increasing the level of insulin and reducing the level of blood glucose. Treatments with both GAG and CsA further decreased the blood glucose level. Moreover, GAG increased both CD4+FoxP3+ and CD8+CD122+PD-1+ Treg numbers in both spleens and lymph nodes of NOD mice. In particular, GAG could reverse a decline in CD4+FoxP3+ Tregs resulted from CsA treatments. The percentage of effector/memory CD8+ T cells (CD44highCD62Llow) was significantly reduced by GAG, especially in the presence of low-doses of CsA. Histopathology also showed that GAG attenuated cellular infiltration and lowered CD3+ T cell numbers around and in islets. Thus, we demonstrated that GAG ameliorated autoimmune T1DM by upregulating both CD4+FoxP3+ and CD8+CD122+PD-1+ Tregs while GAG synergized with CsA to further suppress autoimmunity and T1DM by reversing the decline in CD4+FoxP3+ Tregs resulted from CsA treatments. This study may have important clinical implications for the treatment of T1DM using traditional Chinese medicine.

Cyclovirobuxinum D alleviates cardiac hypertrophy in hyperthyroid rats by preventing apoptosis of cardiac cells and inhibiting the p38 mitogen-activated protein kinase signaling pathway.

OBJECTIVE: To investigate the underlying mechanisms of cyclovirobuxinum D (Cvb-D) on alleviating cardiac hypertrophy in rats. METHODS: Sprague-Dawley rats were randomly divided into 5 groups: control group; levothyroxine-induced cardiac hypertrophy group (model); levothyroxine-induced cardiac hypertrophy + Cvb-D group (Cvb-D); levothyroxine-induced cardiac hypertrophy + captopril group (captopril); levothyroxine-induced cardiac hypertrophy + SB203580 group (SB203580), n=10 for each group. Rats were daily administered the respective drugs continuously for14 days by gastric gavage. A rat model of cardiac hypertrophy was established by intraperitoneal injection of levothyroxine to investigate whether Cvb-D protects against cardiac hypertrophy by inhibiting the p38 mitogen-activated protein kinase (MAPK) signaling pathway and preventing apoptosis of cardiac cells. RESULTS: Treatment with Cvb-D significantly deceased left ventricle hypertrophy, improved the histopathology, hemodynamic conditions, and cardiac function in rats with cardiac hypertrophy. Compared with the normal control group, in rats with cardiac hypertrophy, expression of bax in the heart and phospho-p38 MAPK protein levels were significantly up-regulated (P<0.01 or 0.05), whereas the bcl-2 protein level was down-regulated (P<0.01). In contrast, Cvb-D treatment reversed the changes in bax and phospho-p38 MAPK protein levels but increased the bcl-2 protein level (P<0.01 or 0.05), and these effects were similar to those of captopril and SB203580 (a specific p38MAPK inhibitor) treatment. Furthermore, both Cvb-D, captopril and SB203580 reduced mRNA expression of p38α, p38ß, c-fos, and c-jun mRNA, and Cvb-D had a stronger effect (P<0.01). CONCLUSION: These results demonstrate that Cvb-D protects against cardiac hypertrophy, which is possibly mediated by prevention of cardiac cell apoptosis and inhibition of the p38MAPK signaling pathway.

Zhen-wu-tang attenuates cationic bovine serum albumin-induced inflammatory response in membranous glomerulonephritis rat through inhibiting AGEs/RAGE/NF-κB pathway activation.

Zhen-wu-tang (ZWT), a traditional Chinese compound formula recorded in the Treatise on Febrile Diseases, has significant inhibitory effects on inflammatory damage and oxidative lesions in rats, but its mechanism of action remains unclear. The aim of the present study was to explore whether the anti-inflammatory and anti-oxidative effects of ZWT were mediated by the AGEs/RAGE/NF-κB signaling pathway in rats with cationic bovine serum albumin (C-BSA)-induced membranous glomerulonephritis (MGN). We found that ZWT significantly reduced the production of malondialdehyde (MDA), but enhanced the superoxide dismutase (SOD) activity. The ELISA results showed that ZWT not only reduced the serum levels of AGEs but also decreased the release of inflammatory mediators (TNF-α, IL-1ß, and IL-6). Meanwhile, HE staining showed that pathological kidney injury was alleviated by ZWT. In addition, ZWT suppressed the expression of RAGE1 and NF-κB p65, as well as the nuclear translocation of NF-κB p65. The accumulation of AGEs, oxidative lesions and inflammation damage were reduced by an AGE inhibitor. Thus, the present study demonstrates that AGEs play a role in the pathogenesis of MGN and that AGE inhibition could reduce the inflammatory reactions and oxidative lesions in MGN. In general, ZWT attenuated MGN, in part, by inhibiting the AGEs/RAGE/NF-κB pathway.

Protection Effect of Zhen-Wu-Tang on Adriamycin-Induced Nephrotic Syndrome via Inhibiting Oxidative Lesions and Inflammation Damage.

Zhen-wu-tang (ZWT), a well-known formula in China, is widely used to treat chronic kidney diseases. However, very little information on ZWT's mechanism of action is currently available. In this study, we investigated the possible protective role and underlying mechanism of ZWT on nephrotic syndrome (NS) induced by Adriamycin (intravenous injection, 6.0 mg/kg) in rats using biochemical and histopathological approaches. ZWT decreased urine protein excretion and the serum levels of total cholesterol, triglycerides, blood urea nitrogen, and creatinine significantly in diseased rats. A decrease in plasma levels of total protein and albumin was also recorded in nephropathic rats. Pathological results show an improved pathological state and recovering glomerular structure in ZWT treatment groups. ZWT decreased renal IL-8 level but increased renal IL-4 level. In addition, rats subjected to ZWT exhibited less IgG deposition in glomerulus compared with model group. RT-PCR results showed that ZWT decreased the mRNA expression of NF- κ B p65 and increased the mRNA expression of I κ B. Furthermore, ZWT reduced the level of MDA and increased SOD activity. These results demonstrated that ZWT ameliorated Adriamycin-induced NS in rats possibly by inhibiting Adriamycin-induced inflammation damage, enhancing body's antioxidant capacity, thereby protecting glomerulus from injury.

Qi-Dan Fang ameliorates adriamycin-induced nephrotic syndrome rat model by enhancing renal function and inhibiting podocyte injury.

ETHNOPHARMACOLOGICAL RELEVANCE: Nephrotic syndrome (NS) is a clinical syndrome with a variety of causes, mainly characterized by heavy proteinuria. Podocyte injury plays a key role in proteinuria, one of the principal means for the control of NS is to prevent podocyte injury. Qi-Dan Fang consists of two of the most extensively applied herbal remedies among Traditional Chinese Medicine (TCM) (Radix Astragali Mongolici and Radix Salviae Miltiorrhizae, with a weight ratio of 5:1) which are specifically used for the treatment of various kidney diseases. In previous studies, we found that Qi-Dan Fang provides improvement to patients with adriamycin-induced nephrotic syndrome by alleviating proteinuria and serum lipid. The aim of this study is to study the efficiency of Qi-Dan Fang on NS model rat with renal dysfunction and podocyte injury, something which has not been carried out yet. MATERIALS AND METHODS: The rats were divided into Normal, Model, Jin Gui Shen Qi Pill (4.12 g/kg), Qi-Dan Fang (3.09, 6.17 and 12.34 g/kg/d) groups, they were each given a single tail intravenous injection of Adriamycin (6.0 mg/kg) except for the Normal group and were orally administered dosages of Qi-Dian Fang and Jin Gui Shen Qi pills once daily for 7 weeks. Following the treatment, the content of cystation C (CysC), blood urea nitrogen (BUN), serum creatinine (Scr) were measured with an autobiochemical analyser. The pathomorphological changes to the glomeruli, the mRNA expressions of nephrin, podocin, CD2AP genes and p53, bax, bcl-2 proteins expressions were also carried out to probe the effects of Qi-Dan Fang. RESULTS: (1) Qi-Dan Fang treatment raised the level of CysC in blood serum while lowering the content of BUN and Scr in the adriamycin-induced nephrotic syndrome rat model; (2) Long-term administration of Qi-Dan Fang was able to ameliorate pathomorphological change of glomeruli and repair the organization structure of Glomerulus; (3) Qi-Dan Fang could increase the mRNA expression of nephrin, podocin and CD2AP genes, down-regulate the expression of p53, bax proteins, while increased bcl-2 protein to protect the podocyte and restore Glomerular selective filtration function. CONCLUSIONS: Results of our present studies reveal that Qi-Dan Fang is able to enhance renal function, inhibit podocyte injury to provide improvements to the Adriamycin-induced nephrotic syndrome.