Recombinant bivalent subunit vaccine combining truncated VP4 from P[7] and P[23] induces protective immunity against prevalent porcine rotaviruses.

Porcine rotaviruses (PoRVs) cause severe economic losses in the swine industry. P[7] and P[23] are the predominant genotypes circulating on farms, but no vaccine is yet available. Here, we developed a bivalent subunit PoRV vaccine using truncated versions (VP4*) of the VP4 proteins from P[7] and P[23]. The vaccination of mice with the bivalent subunit vaccine elicited more robust neutralizing antibodies (NAbs) and cellular immune responses than its components, even at high doses. The bivalent subunit vaccine and inactivated bivalent vaccine prepared from strains PoRVs G9P[7] and G9P[23] were used to examine their protective efficacy in sows and suckling piglets after passive immunization. The immunized sows showed significantly elevated NAbs in the serum and colostrum, and the suckling piglets acquired high levels of sIgA antibodies from the colostrum. Challenging subunit-vaccinated or inactivated-vaccinated piglets with homologous virulent strains did not induce diarrhea, except in one or two piglets, which had mild diarrhea. Immunization with the bivalent subunit vaccine and inactivated vaccine also alleviated the microscopic lesions in the intestinal tissues caused by the challenge with the corresponding homologous virulent strain. However, all the piglets in the challenged group displayed mild to watery diarrhea and high levels of viral shedding, whereas the feces and intestines of the piglets in the bivalent subunit vaccine and inactivated vaccine groups had lower viral loads. In summary, our data show for the first time that a bivalent subunit vaccine combining VP4*P[7] and VP4*P[23] effectively protects piglets against the diarrhea caused by homologous virulent strains.IMPORTANCEPoRVs are the main causes of diarrhea in piglets worldwide. The multisegmented genome of PoRVs allows the reassortment of VP4 and VP7 genes from different RV species and strains. The P[7] and P[23] are the predominant genotypes circulating in pig farms, but no vaccine is available at present in China. Subunit vaccines, as nonreplicating vaccines, are an option to cope with variable genotypes. Here, we have developed a bivalent subunit candidate vaccine based on a truncated VP4 protein, which induced robust humoral and cellular immune responses and protected piglets against challenge with homologous PoRV. It also appears to be safe. These data show that the truncated VP4-protein-based subunit vaccine is a promising candidate for the prevention of PoRV diarrhea.

Isolation and characterization of a G9P[23] porcine rotavirus strain AHFY2022 in China.

Rotavirus group A (RVA) is a main pathogen causing diarrheal diseases in humans and animals. Various genotypes are prevalent in the Chinese pig herd. The genetic diversity of RVA lead to distinctly characteristics. In the present study, a porcine RVA strain, named AHFY2022, was successfully isolated from the small intestine tissue of piglets with severe diarrhea. The AHFY2022 strain was identified by cytopathic effects (CPE) observation, indirect immunofluorescence assay (IFA), electron microscopy (EM), high-throughput sequencing, and pathogenesis to piglets. The genomic investigation using NGS data revealed that AHFY2022 exhibited the genotypes G9-P[23]-I5-R1-C1-M1-A8-N1-T1-E1-H1, using the online platform the Bacterial and Viral Bioinformatics Resource Center (BV-BRC) (https://www.bv-brc.org/). Moreover, experimental inoculation in 5-day-old and 27-day-old piglets demonstrated that AHFY2022 caused severe diarrhea, fecal shedding, small intestinal villi damage, and colonization in all challenged piglets. Taken together, our results detailed the virological features of the porcine rotavirus G9P[23] from China, including the whole-genome sequences, genotypes, growth kinetics in MA104 cells and the pathogenicity in suckling piglets.

Combining Multifunctional Delivery System with Blood-Brain Barrier Reversible Opening Strategy for the Enhanced Treatment of Alzheimer's Disease.

Alzheimer's disease (AD) is a neurodegenerative illness characterized by intracellular tau-phosphorylation, ß-amyloid (Aß) plaques accumulation, neuroinflammation, and impaired behavioral ability. Owing to the lack of effective brain delivery approaches and the presence of the blood-brain barrier (BBB), current AD therapeutic endeavors are severely limited. Herein, a multifunctional delivery system (RVG-DDQ/PDP@siBACE1) is elaborately combined with a protein kinase B (AKT) agonist (SC79) for facilitating RVG-DDQ/PDP@siBACE1 to target and penetrate BBB, enter brain parenchyma, and further accumulate in AD brain lesion. Moreover, compared with the unitary dose of RVG-DDQ/PDP@siBACE1, this collaborative therapy strategy exhibits a distinctive synergistic function including scavenging reactive oxygen species (ROS), decreasing of Aß production, alleviating neuroinflammation by promoting the polarized microglia into the anti-inflammatory M2-like phenotype and significantly enhancing the cognitive functions of AD mice. More strikingly, according to these results, an innovative signaling pathway "lncRNA MALAT1/miR-181c/BCL2L11" is found that can mediate the neuronal apoptosis of AD. Taken together, combining the brain targeted delivery system with noninvasive BBB opening can provide a promising strategy and platform for targeting treatment of AD and other neurodegenerative diseases.

Astragaloside IV Inhibited Podocyte Pyroptosis in Diabetic Kidney Disease by Regulating SIRT6/HIF-1α Axis.

To investigate the effect of astragaloside IV (AS) on podocytes pyroptosis in diabetic kidney disease (DKD). Forty male Sprague-Dawley rats were randomly divided into normal group (n = 10) and model group (n = 30). Rats in model group were intraperitoneally injected streptozotocin (60 mg/kg) for 3 days to induce DKD. Then rats were divided into DKD group, AS group, and UBCS group. The AS group was given 40 mg/kg/d of AS by gavage, and UBCS group was given 50 mg/kg/d of UBCS039 by gavage, and normal group and DKD group were given the same amount saline for 8 weeks, once a day. Hematoxylin-eosin and masson staining were used to observe pathology of kidney. Rat podocytes were divided into normal group, mannitol hypertonic group, high-glucose group, UBCS group, OSS group, and AS group. Western blotting, quantitative real-time polymerase chain reaction, immunofluorescence, and flow cytometry were used to analyze pyroptosis-related markers and reactive oxygen species (ROS) levels. Results showed that AS inhibited ROS and alleviated podocytes pyroptosis in rats by increasing expression of sirtuin 6 (SIRT6) and decreasing expression of hypoxia inducible factor 1 subunit alpha (HIF-1α). UBCS039 and AS enhanced SIRT6 level, decreased HIF-1α level, and finally improved pyroptosis of podocytes in vitro, whereas OSS-128167 showed the opposite effect for podocytes pyroptosis. AS improved podocytes pyroptosis in DKD by regulating SIRT6/HIF-1α pathway, thereby alleviating injury of DKD.

Normalizing Tumor Blood Vessels to Improve Chemotherapy and Inhibit Breast Cancer Metastasis by Multifunctional Nanoparticles.

The abnormal tumor blood vessels with high leakage can promote tumor cells to infiltrate into the systemic circulation and increase the risk of tumor metastasis. In addition, chemotherapy may destroy tumor blood vessels and further aggravate metastasis. Normalizing tumor blood vessels can reduce vascular leakage and increase vascular integrity. The simultaneous administration of vascular normalization drugs and chemotherapy drugs may resist the blood vessels' destruction of chemotherapy. Here, multifunctional nanoparticles (CCM@LMSN/DOX&St), which combined chemotherapy with tumor blood vessel normalization, were prepared for the treatment of breast cancer. The results showed that CCM@LMSN/DOX&St-loaded sunitinib (St) promoted the expression of junction proteins Claudin-4 and VE-cadherin of endothelial cells, reversed the destruction of DOX to the endothelial cell layer, protected the integrity of the endothelial cell layer, and inhibited the migration of 4T1 tumor cells across the endothelial cell layer. In vivo experiments showed that CCM@LMSN/DOX&St effectively inhibited tumor growth in situ; what is exciting was that it also inhibited distal metastasis of breast cancer. CCM@LMSN/DOX&St encapsulated with St can normalize tumor blood vessels, reverse the damage of DOX to tumor blood vessels, increase the integrity of blood vessels, and prevent tumor cell invasion into blood vessels, which can inhibit breast cancer spontaneous metastasis and reduce chemotherapy-induced metastasis. This drug delivery platform effectively inhibited the progression of tumors and provided a promising solution for effective tumor treatment.

GSH-Responsive Polymeric Micelles for Remodeling the Tumor Microenvironment to Improve Chemotherapy and Inhibit Metastasis in Breast Cancer.

The tumor microenvironment (TME) of breast cancer is hypoxic, which can promote tumor progression, including invasion and metastasis, and limit the efficacy of anti-tumor treatment. Nitric oxide (NO) can dilate blood vessels, effectively alleviate hypoxia, and regulate the TME, which has the potential to improve the anti-tumor therapeutic efficacy. Here, chitosan (CO) and octadecylamine (ODA) were linked by the disulfide bond, and the LinTT1 peptide was linked onto CO-SS-ODA for targeting tumor cells and endothelial cells in tumors. The NO donor S-nitroso-N-acetylpenicillamine (SNAP) was connected to CO. Doxorubicin (DOX) was encapsulated, and GSH hierarchically responsive polymer micelles (TSCO-SS-ODA/DOX) were constructed for the treatment of breast cancer. The micelles had differently responsive drug release in different GSH concentrations. In endothelial cells, the micelles rapidly responded to release NO. In tumor cells, the disulfide bond rapidly broke and released DOX to effectively kill tumor cells. The disulfide bond was not sensitive to GSH concentration in endothelial cells, which had less release of DOX. The killing effect of the micelles to endothelial cells was much lower than that to tumor cells. The cell selective drug release of the drug delivery systems enabled safe and effective treatment of drugs. TSCO-SS-ODA/DOX, which had the excellent ability to target tumors, can alleviate tumor hypoxia, decrease the infiltration of M2 macrophages in tumors, increase the infiltration of M1 macrophages in tumors, and remodel the TME. Notably, TSCO-SS-ODA/DOX can significantly inhibit the growth of the primary tumor and effectively inhibit tumor metastasis. The drug delivery system provided a potential solution for effectively treating breast cancer.

Aucubin alleviates methotrexate-induced enteritis in rats by inducing autophagy.

One of the toxic side effects of methotrexate (MTX) is enteritis. Aucubin, an iridoid glycoside derived from traditional medicinal herbs, has been proven to have anti-inflammation, anti-apoptosis and anti-oxidation properties. This work explored the effect and mechanism of aucubin in treating MTX-induced enteritis in a rat model. Two doses of aucubin (5 and 10 mg/kg) were adopted for the assessment of its pharmacological activity. We observed that in rats with MTX-induced enteritis, the body weight and small intestinal weight decreased. The intestine barrier was injured, as reflected by pathological examinations and an increase in D-lactate and diamine oxidase concentration in serum. Intestinal inflammation was shown by the observation of macrophages in the intestine and the concentrations of inflammatory cytokines tumour necrosis factor-α (TNF-α) and interleukin-6 (IL-6) in serum. The NLR family pyrin domain containing 3 (NLRP3) inflammasome was shown to be activated by the enhancement of NLRP3, cleaved-caspase 1, IL-18 and IL-1ß. Moreover, autophagy was reflected by transmission electron microscopy as slightly induced, along with changes in autophagy-related markers microtubule-associated protein 1 light chain 3 (LC3) and Beclin1. Remarkably, aucubin treatment attenuated the MTX-induced disease activity index increase, intestinal damage, inflammatory response and NLRP3 inflammasome activation, but provoked autophagy. Rapamycin, an autophagy activator, showed similar therapeutic effects to aucubin on MTX-induced enteritis. However, 3-methyladenine, an autophagy inhibitor, reversed the protective effects of aucubin. These findings prompted the hypothesis that aucubin alleviates MTX-induced enteritis by aggravating autophagy. This study might provide evidence for further investigation on the therapeutic role of aucubin in MTX-resulted enteritis.

Enhanced nitrogen removal via Yarrowia lipolytica-mediated nitrogen and related metabolism of Chlorella pyrenoidosa from wastewater.

We investigated the optimum co-culture ratio with the highest biological nitrogen removal rate, revealing that chemical oxygen demand, total nitrogen (TN), and ammoniacal nitrogen (NH3-N) removal was increased in the Chlorella pyrenoidosa and Yarrowia lipolytica co-culture system at a 3:1 ratio. Compared with the control, TN and NH3-N content in the co-incubated system was decreased within 2-6 days. We investigated mRNA/microRNA (miRNA) expression in the C. pyrenoidosa and Y. lipolytica co-culture after 3 and 5 days, identifying 9885 and 3976 differentially expressed genes (DEGs), respectively. Sixty-five DEGs were associated with Y. lipolytica nitrogen, amino acid, photosynthetic, and carbon metabolism after 3 days. Eleven differentially expressed miRNAs were discovered after 3 days, of which two were differentially expressed and their target mRNA expressions negatively correlated with each other. One of these miRNAs regulates gene expression of cysteine dioxygenase, hypothetical protein, and histone-lysine N-methyltransferase SETD1, thereby reducing amino acid metabolic capacity; the other miRNA may promote upregulation of genes encoding the ATP-binding cassette, subfamily C (CFTR/MRP), member 10 (ABCC10), thereby promoting nitrogen and carbon transport in C. pyrenoidosa. These miRNAs may further contribute to the activation of target mRNAs. miRNA/mRNA expression profiles confirmed the synergistic effects of a co-culture system on pollutant disposal.

Dietary calcium and nonphosphate phosphorus interaction influences tibiotarsus development and related gene expression of broilers from 1 to 21 days of age.

The dietary needs of calcium (Ca) and phosphorus (P) are interdependent, thus accurate evaluation of Ca and P requirements of broilers to support skeleton health and optimal growth is critical. The present study was carried out to investigate the effects of dietary Ca and nonphytate P (NPP) levels and their interactions on growth performance, tibiotarsus characteristics, tibiotarsus metabolism-related enzyme and proteins, and their gene expression of broilers, so as to provide a rational recommendation for Ca and NPP levels in diet. A total of 540 one-day-old Arbor Acres male broilers were randomly allotted to 1 of 15 treatments with 6 replicate cages of 6 birds per cage for each treatment in a completely randomized design involving a 5 × 3 factorial arrangement of treatments (5 levels of Ca × 3 levels of NPP). The birds were fed the corn-soybean meal diet containing 0.60%, 0.70%, 0.80%, 0.90%, or 1.00% Ca and 0.35%, 0.40%, or 0.45% NPP for 21 d. Dietary Ca level affected (P < 0.03) the bone mineral density, bone mineral content (BMC), breaking strength, ash percentage and ash Ca contents in tibia, which showed linear (P < 0.006) responses to dietary Ca levels. Dietary NPP level affected (P < 0.05) tibia BMC, ash percentage, and FGF23 mRNA level. Broilers that received 0.40% and 0.45% NPP had higher (P < 0.04) tibia BMC and ash percentage than those that received 0.35% NPP, but no differences (P > 0.05) were found between 0.40% and 0.45% NPP. Broilers that received 0.40% NPP had higher (P = 0.02) tibia FGF23 mRNA level than those that received 0.35% NPP, but no differences (P > 0.05) were detected between 0.40% and 0.45% NPP or 0.45% and 0.35% NPP. The interactions between dietary Ca and NPP affected (P < 0.05) ADG, ALP activity, bone gal protein, FGF23 contents, and the mRNA expression levels ALP and bone gal protein in tibia of broilers. Results from the present study indicate that dietary Ca and NPP interaction influences growth, tibiotarsus development, and related gene expression of broiler chickens. Considering all the criteria, the dietary levels of 0.90% Ca and 0.45% NPP would be optimal for both growth and tibiotarsus development of broilers fed a conventional corn-soybean meal diet from 1 to 21 d of age.

Enhanced brain delivery of hypoxia-sensitive liposomes by hydroxyurea for rescue therapy of hyperacute ischemic stroke.

Ischemic stroke is characterized by high morbidity, disability, and mortality. Unfortunately, the only FDA-approved pharmacological thrombolytic, alteplase, has a narrow therapeutic window of only 4.5 h. Other drugs like neuroprotective agents have not been clinically used because of their low efficacy. To improve the efficacy of neuroprotective agents and the effectiveness of rescue therapies for hyperacute ischemic stroke, we investigated and verified the variation trends of the blood-brain barrier (BBB) permeability and regional cerebral blood flow over 24 h in rats that had ischemic strokes. Hypoperfusion and the biphasic increase of BBB permeability are still the main limiting factors for lesion-specific drug distribution and drug brain penetration. Herein, the nitric oxide donor hydroxyurea (HYD) was reported to downregulate the expression of tight junction proteins and upregulate intracellular nitric oxide content in the brain microvascular endothelial cells subjected to oxygen-glucose deprivation, which was shown to facilitate the transport of liposomes across  brain endothelial monolayer in an in vitro model. HYD also increased the BBB permeability and promoted microcirculation in the hyperacute phase of stroke. The neutrophil-like cell-membrane-fusogenic hypoxia-sensitive liposomes exhibited excellent performance in targeting the inflamed brain microvascular endothelial cells, enhancing cell association, and promoting rapid hypoxic-responsive release in the hypoxic microenvironment. Overall, the combined HYD and hypoxia-sensitive liposome dosing regimen effectively decreased the cerebral infarction volume and relieved neurological dysfunction in rats that had ischemic strokes; these therapies were involved in the anti-oxidative stress effect and the neurotrophic effect mediated by macrophage migration inhibitory factor.

Adult upper-limb forearm lymphoma hygroma (cystic lymphangioma): A case report.

INTRODUCTION: Lymphoma hygroma (LH) that is the most common type of lymphangioma, but it rarely occurs in the forearm. It may show localized invasive behavior, but is benign. CASE PRESENTATION: A 42-year-old woman presented to our hospital with a growing strip-like mass in the right forearm that had been detected 3 years earlier. Ultrasound examination showed a subcutaneous strip of low back vocal cords on the right forearm. Further magnetic resonance imaging (MRI) showed irregular strip-like dilated lymphatic vessels characteristic of LH with low T1 signal intensity and high T2 signal intensity. After radical surgical resection, hematoxylin-eosin (H & E) and immunohistochemical (IHC) staining of cystic LH endothelial cells labeled with monoclonal antibody D2-40 showed a dilated lymphangioma with no evidence of malignancy. After 7 months of follow-up, no tumor recurrence was seen and the effect was satisfactory. CLINICAL DISCUSSION: A combination of previous trauma history, signs and symptoms, and imaging evaluation are necessary to provide clues to LH, but the final diagnosis is likely to be made by pathologic evaluation of the resected specimen. Although there are many treatment modalities, all also have different outcomes. The absence of complete resection resulting in a tumor remnant is the foremost cause of LH recurrence, so we believe that the preferred approach against LH remains complete surgical resection. CONCLUSIONS: LH is benign and generally asymptomatic lesions with mild bio-behavior. As there are occasional confusing presentations, similar cystic lesions should still be considered with caution for the disease. Although MRI provides superior advantages for its diagnosis, the confirmation of diagnosis still requires histological examination. Radical lesion resection is a very safe and effective option for the treatment of LH.

Corynoline ameliorates dextran sulfate sodium-induced colitis in mice by modulating Nrf2/NF-κB pathway.

OBJECTIVE: Corynoline is an active substance extracted from Corydalis bungeana Turcz and exerts a therapeutic effect in multiple diseases by alleviating inflammatory response. The present study sought to elucidate the role of corynoline in ulcerative colitis (UC). METHODS: The experimental colitis models were induced in BALB/c mice via receiving a drinking water supplemented with 3.5% (I) dextran sulfate sodium (DSS) ad libitum for 7 days. RESULTS: Corynoline administration inhibited body weight loss, colon shortening, disease activity index and colonic pathomorphological changes in DSS-treated mice. Besides, corynoline down-regulated the levels of pro-inflammatory interleukin (IL)-1ß, IL-6 and tumor necrosis factor Alpha (TNF-α), as well as decreased myeloperoxidase (MPO) activity in the colon of DSS-treated mice. In addition, severe oxidative stress in the colonic tissues of DSS-treated was mitigated by corynoline treatment. However, these beneficial effects were reversed by a specific nuclear factor E2-related factor 2 (Nrf2) inhibitor ML385 intervention. Further evidence confirmed that corynoline promoted Nrf2 nuclear migration and heme oxygenase-1 gene expression in the colonic tissues of UC mice. Besides, corynoline treatment restrained colonic nuclear factor-kappa B (NF-κB) activation as proved by the decrease in phosphorylation and nuclear translocation of NF-κB. CONCLUSIONS: Corynoline ameliorates DSS-induced mouse colitis, which may provide a promising therapeutic strategy for UC treatment.

Case Report: A case of uterine leiomyosarcoma metastasized to the vena cava, excised with the aid of preoperative CT three-dimensional imaging.

Leiomyosarcoma of the uterus (ULMS) is a rare malignant tumor originating from embryonic mesenchymal cells. ULMS tends to metastasize to the lungs, lymph nodes, liver, and bone. Computed tomography three-dimensional (CT 3D) imaging is an advanced diagnostic technique that can track the vessels and their relationships with tumors and reveal the invasion of vessels, including small vessels, around tumors in any slice. Here, we describe a case in which ULMS extended to the retrohepatic inferior vena cava. To date, no report has described resection of metastatic ULMS of the vena cava through supplemental CT 3D imaging. Our patient presented with right lumbar abdominal pain as the main symptom. After using CT 3D reconstruction to accurately assess the relationship between the tumor and the surrounding organs and blood vessels before the operation, the operation was successfully completed through multidisciplinary surgical collaboration.

Cell membrane coated-nanoparticles for cancer immunotherapy.

Cancer immunotherapy can effectively inhibit cancer progression by activating the autoimmune system, with low toxicity and high effectiveness. Some of cancer immunotherapy had positive effects on clinical cancer treatment. However, cancer immunotherapy is still restricted by cancer heterogeneity, immune cell disability, tumor immunosuppressive microenvironment and systemic immune toxicity. Cell membrane-coated nanoparticles (CMCNs) inherit abundant source cell-relevant functions, including "self" markers, cross-talking with the immune system, biological targeting, and homing to specific regions. These enable them to possess preferred characteristics, including better biological compatibility, weak immunogenicity, immune escaping, a prolonged circulation, and tumor targeting. Therefore, they are applied to precisely deliver drugs and promote the effect of cancer immunotherapy. In the review, we summarize the latest researches of biomimetic CMCNs for cancer immunotherapy, outline the existing specific cancer immune therapies, explore the unique functions and molecular mechanisms of various cell membrane-coated nanoparticles, and analyze the challenges which CMCNs face in clinical translation.

Metabolic utilization of intravenously injected iron from different iron sources in target tissues of broiler chickens.

No information is available regarding the utilization of iron (Fe) from different Fe sources at a target tissue level. To detect differences in Fe metabolic utilization among Fe sources, the effect of intravenously injected Fe on growth performance, hematological indices, tissue Fe concentrations and Fe-containing enzyme activities and gene expressions of Fe-containing enzymes or protein in broilers was investigated. On d 22 post-hatching, a total of 432 male chickens were randomly allotted to 1 of 9 treatments in a completely randomized design. Chickens were injected with either a 0.9% (wt/vol) NaCl solution (control) or a 0.9% NaCl solution supplemented with Fe sulphate or 1 of 3 organic Fe sources. The 3 organic Fe sources were Fe chelates with weak (Fe-MetW), moderate (Fe-ProtM) or extremely strong (Fe-ProtES) chelation strength. The 2 Fe dosages were calculated according to the Fe absorbabilities of 10% and 20% every 2 d for a duration of 20 d. Iron injection did not affect (P > 0.05) ADFI, ADG or FCR during either 1 to 10 d or 11 to 20 d after injections. Hematocrit and Fe concentrations in the liver and kidney on d 10 after Fe injections, and Fe concentrations in the liver or pancreas and ferritin heavy-chain (FTH1) protein expression level in the liver or spleen on d 20 after Fe injections increased (P ≤ 0.05) as injected Fe dosages increased. When the injected Fe level was high at 20% Fe absorbability, the chickens injected with Fe-ProtES had lower (P < 0.001) liver or kidney Fe concentrations and spleen FTH1 protein levels than those injected with Fe-MetW or Fe-ProtM on d 20 after injections. And they had lower (P < 0.05) liver cytochrome C oxidase mRNA levels on d 20 after injections than those injected with Fe-MetW or Fe sulphate. The results from this study indicate that intravenously injected Fe from Fe-ProtES was the least utilizable and functioned in the sensitive target tissue less effectively than Fe from Fe sulfate, Fe-MetW or Fe-ProtM.

Potential Molecular Mechanism of Yishen Capsule in the Treatment of Diabetic Nephropathy Based on Network Pharmacology and Molecular Docking.

Purpose: Using network pharmacology and molecular docking to explore the mechanism of Yishen Capsule in the treatment of diabetic nephropathy. Materials and Methods: Active components of Yishen Capsule were obtained using database such as TCMSP and TCMID. UniProt protein database was used to screen and standardize the human-derived targets of the active chemical components. Diabetic nephropathy (DN) targets were obtained from databases such as GeneCards, OMIM, TTD, DisGeNET and DrugBank. A network of "Yishen Capsule Components-diabetic nephropathy Targets-Pathways" was constructed by analyzing data above to screening out core targets for molecular docking verification. DN is induced by streptozocin in rats after left nephrectomy. Renal tubular epithelial cells (RTECs) was isolated  and cultured under high glucose conditions. Based on these experimental models, key pathway target genes screened by network pharmacology were verified both in vitro and in vivo. Results: The main active components of Yishen Capsule in the treatment of DN include quercetin, kaempferol, gallic acid, astragaloside IV, etc. Some key targets (such as AR, AKT1, TP53, ESR1, JUN) and important signal pathways (such as AGE-RAGE, HIF-1 and JAK-STAT signal pathway) were included in the treatment of DN with Yishen Capsule. Molecular docking assay showed that most of the targets have good binding activity with the components of Yishen Capsule. Based on the results of network pharmacology, key target proteins in HIF-1α and JAK2/STAT3 signaling pathways were selected for experimental verification. Results presented that HIF-1α, JAK2, STAT3, TGF-ß and MCP-1 were increased under high glucose environment. With the treatment of Yishen Capsule, the expression of HIF-1α further increased, while the expression of JAK2, STAT3, MCP-1 and TGF-ß was decreased. Conclusion: This study revealed the mechanism of Yishen Capsule in the treatment of DN, which possesses the characteristics of multi-component, multi-target, and multi-pathway. Further experiments confirmed that Yishen Capsule interfered with HIF-1α and JAK/STAT signaling pathways to reduce inflammation and fibrosis damage in the kidney tissue of rats with diabetic nephropathy.

Evaluation of optimal dietary calcium level by bone characteristics and calcium metabolism-related gene expression of broilers from 22 to 42 d of age.

The current dietary Ca recommendation of broilers is primarily based on the previous studies carried out more than 30 yr ago. However, the modern commercial broilers are quite different from those more than 30 yr ago. The present experiment was conducted to evaluate an optimal dietary Ca level by bone characteristics and Ca metabolism-related gene expression of broilers fed a corn-soybean meal diet from 22 to 42 d of age. A total of 252 22-d-old Arbor Acres male broilers were randomly assigned to 1 of 7 treatments with 6 replicate cages of 6 birds per cage for each treatment. Broilers were fed the corn-soybean meal diets containing 0.50%, 0.60%, 0.70%, 0.80%, 0.90%, 1.00%, or 1.10% Ca for 21 d, and each diet contained 0.31% non-phytate P. The results showed that the mineral contents in tibia and middle toe bone, mineral density in tibia and middle toe bone, middle toe ash percentage, middle toe ash Ca percentage, and tibia alkaline phosphatase mRNA expression level of broilers were influenced (P < 0.04) by dietary Ca level and increased quadratically (P < 0.05) as dietary Ca level increased. The estimates of optimal dietary Ca levels were 0.55%, 0.60%, 0.70%, 0.72%, 0.63%, 0.66%, and 0.70%, respectively, based on the best fitted broken-line, quadratic, or asymptotic models (P < 0.02) of the above sensitive indices. These results indicate that the optimal dietary Ca level would be 0.72% to support all of the Ca metabolism and bone development of broilers fed the corn-soybean meal diet from 22 to 42 d of age.

The present experiment was conducted to evaluate an optimal dietary Ca level by bone characteristics and Ca metabolism-related gene expression of broilers fed a corn-soybean meal diet from 22 to 42 d of age. A total of 252 22-d-old Arbor Acres male broilers were randomly assigned to 1 of 7 treatments with 6 replicate cages of 6 birds per cage for each treatment. Broilers were fed the corn-soybean meal diets containing 0.50%, 0.60%, 0.70%, 0.80%, 0.90%, 1.00%, or 1.10% Ca for 21 d, and each diet contained 0.31% non-phytate P. The results showed that the tibia and middle toe bone mineral contents, tibia and middle toe bone mineral density, middle toe ash percentage, middle toe ash Ca percentage, and tibia alkaline phosphatase mRNA expression level were sensitive criteria to estimate the optimal dietary Ca levels of broilers. The estimates of optimal dietary Ca levels were 0.55% to 0.72% based on the above sensitive criteria. These results indicate that the optimal dietary Ca level would be 0.72% to support all of the Ca metabolism and bone development of broilers fed the corn-soybean meal diet from 22 to 42 d of age.

Dietary calcium requirements of broilers fed a conventional corn-soybean meal diet from 1 to 21 days of age.

BACKGROUND: The current calcium (Ca) recommendation for broilers is primarily based on studies conducted more than 30 years ago with birds of markedly different productive potentials from those which exist today. And the response indicators in these studies are mainly growth performance and bone ash percentage. Therefore, the present study was carried out to investigate the effect of dietary Ca level on growth performance, serum parameters, bone characteristics and Ca metabolism-related gene expressions, so as to estimate dietary Ca requirements of broilers fed a conventional corn-soybean meal diet from 1 to 21 days of age. METHODS: A total of 420 1-day-old Arbor Acres male broilers were randomly assigned to 1 of 7 treatments with 6 replicates (10 birds per cage) and fed the corn-soybean meal diets containing 0.60%, 0.70%, 0.80%, 0.90%, 1.00%, 1.10% or 1.20% Ca for 21 days. Each diet contained the constant non-phytate phosphorus content of about 0.39%. RESULTS: The average daily gain decreased linearly (P < 0.001) as dietary Ca level increased. The serum and tibia alkaline phosphatase (ALP) activities, tibia bone mineral density (BMD), middle toe BMD, tibia ash percentage, tibia breaking strength, and tibia ALP protein expression level were affected (P < 0.05) by dietary Ca level, and showed significant quadratic responses (P < 0.02) to dietary Ca levels. The estimates of dietary Ca requirements were 0.80 to 1.00% based on the best fitted broken-line or quadratic models (P < 0.03) of the above serum and bone parameters, respectively. CONCLUSIONS: The results from the present study indicate that the Ca requirements would be about 0.60% to obtain the best growth rate, and 1.00% to meet all of the Ca metabolisms and bone development of broilers fed a conventional corn-soybean meal diet from 1 to 21 days of age.

Determination of optimal dietary selenium levels by full expression of selenoproteins in various tissues of broilers from 22 to 42 d of age.

The current NRC dietary selenium (Se) requirement (0.15 mg/kg) of broilers from 22 to 42 d of age is primarily based on a previous study reported in 1986, which might not be applicable to modern classes of rapidly growing broilers. The present experiment was conducted to determine the optimal dietary Se level for meeting metabolic and functional Se requirements of broilers fed a corn-soybean meal diet from 22 to 42 d of age. A total of 336 Arbor Acres male broilers at 22 d old were randomly assigned to 1 of 6 treatments with 7 replicates and fed a basal corn-soybean meal diet (control, containing 0.014 mg Se/kg) and the basal diet supplemented with 0.10, 0.20, 0.30, 0.40, or 0.50 mg Se/kg from Na2SeO3 for 21 d. The results showed that the Se concentrations in plasma, liver, kidney, pancreas, breast and thigh muscles, the activity of glutathione peroxidase (GPX) in plasma, liver and kidney, the mRNA expression levels of Gpx4, selenoprotein (Seleno) h and Selenou in liver, S elenop and Selenoh in kidney, and the protein expression levels of GPX4 in the liver and kidney of broilers were affected (P < 0.05) by supplemental Se level, and increased quadratically (P < 0.05) with the increase of supplemental Se level. The estimates of optimal dietary Se levels were 0.10 to 0.49 mg/kg based on the fitted broken-line or asymptotic models (P < 0.0001) of the above Se concentration indices, and 0.08 to 0.37 mg/kg based on the fitted broken-line, quadratic or asymptotic models (P < 0.007) of the above selenoprotein expression indices. These results indicate that the optimal dietary Se levels would be 0.49 mg/kg to support the maximum Se concentrations and 0.37 mg/kg to support the full expression of selenoproteins in plasma and various tissues of broilers fed a corn-soybean meal diet from 22 to 42 d of age.

[Effects of astragaloside IV on delaying kidney aging and its mechanisms].

OBJECTIVE: To investigate the mechanisms of Astragaloside Ⅳ on inhibiting apoptosis and delaying kidney aging in rats by regulating SIRT1/p53 signaling pathway. METHODS: The aging model was established by subcutaneous injection of D-galactose 200 mg/(kg·d). SPF-grade healthy male SD rats were randomly divided into 4 groups: normal control group (intragastric infusion of 5 ml/(kg·d) normal saline), aging model group (intragastric infusion of 5 ml/(kg·d) normal saline), Astragaloside IV group (intragastric infusion of 40 mg/(kg·d) Astragaloside IV),and SRT1720 group( intragastric infusion of 20 mg/(kg·d) SRT1720), with 10 rats in each group. After 8 weeks, the serum samples of rats were collected to detect the levels of renal function (creatinine and urea nitrogen) and senescent associated secretory phenotype (TGF-ß and IL-6) by ELISA. The renal tissues of rats were obtained for HE and Masson staining. The protein and mRNA expressions of SIRT1, p53, Bcl-2, Bax, p21 and pRb were detected by Western blot and RT-PCR. RESULTS: Serum creatinine and urea nitrogen levels in the aging model group were higher than those in the normal group, but there was no significant difference in each group (P＞0.05). The serum levels of TGF-ß and IL-6 in the aging model group were higher than those in the normal group (P＜0.05), and which in the Astragaloside IV group and SRT1720 group were lower than those in the model group (P＜0.05). There was no significant differences between Astragaloside IV group and SRT1720 group (P＞0.05). The results of pathological staining of renal tissues showed that, compared with the normal group, the renal tubules dilated, local atrophy, infiltration of inflammatory cells and proliferation of collagen fibers were observed in the aging model group. Compared with the aging model group, the pathological changes were alleviated in Astragaloside IV group and SRT1720 group. The results of Western blot and RT-PCR showed that, compared with the normal group, the protein and mRNA expressions of SIRT1 and pRb in the renal tissue of the aging group were decreased, the protein expression of Bcl-2 was decreased(P＜0.05), and the protein and mRNA expressions of p53 and p21 were increased, the protein expression of Bax was increased(P＜0.05). Compared with the aging group, Astragaloside IV and SRT1720 improved the above-mentioned indexes (P＜0.05). CONCLUSION: Astragaloside IV can delay kidney aging by regulating the SIRT1/p53 signaling pathway.