Study on the immune-enhancing and inhabiting transmissible gastroenteritis virus effects of polysaccharides from Cimicifuga rhizoma.

Cimicifugae rhizoma is a traditional Chinese herbal medicine in China, and modern pharmacological research showed that it has obvious antiviral activity. Many polysaccharides have been proved to have immune enhancement and antiviral activity, but there are few studies on the biological activity of Cimicifuga rhizoma polysaccharide (CRP). The aim was to explore the character of CRP and its effects on improving immune activity and inhibiting transmissible gastroenteritis virus (TGEV). The monosaccharide composition, molecular weight, fourier transform infrared spectra and electron microscopy analysis of CRP was measured. The effect of CRP on immune activity in lymphocytes and RAW264.7 cells were studied by colorimetry, FITC-OVA fluorescent staining and ELISA. The effect of CRP on TGEV-infected PK-15 cells was determined using Real-time PCR, Hoechst fluorescence staining, trypan blue staining, acridine orange staining, Annexin V-FITC/PI fluorescent staining, DCFH-DA loading probe, and JC-1 staining. Network pharmacology was used to predict the targets of CRP in enhancing immunity and anti-TGEV, and molecular docking was used to further analyze the binding mode between CPR and core targets. The results showed that CRP was mainly composed of glucose and galactose, and its molecular weight was 64.28 kDa. The content of iNOS and NO in CRP group were significantly higher than the control group. CRP (125 and 62.5 µg/mL) could significantly enhance the phagocytic capacity of RAW264.7 cells, and imprive the content of IL-1ß content compared with control group. 250 µg/mL of CRP possessed the significant inhibitory effect on TGEV, which could significantly reduce the apoptosis compared to TGVE group and inhibit the decrease in mitochondrial membrane potential compared to TGVE group. The mRNA expression of TGEV N gene in CRP groups was significantly lower than TGEV group. PPI showed that the core targets of immune-enhancing were AKT1, MMP9, HSP90AA1, etc., and the core targets of TGE were CASP3, MMP9, EGFR, etc. Molecular docking show that CRP has binding potential with target. These results indicated that CRP possessed the better immune enhancement effect and anti-TGEV activity.

Hypoxia activation attenuates progesterone synthesis in goat trophoblast cells via NR1D1 inhibition of StAR expression†.

Trophoblast plays a crucial role in gestation maintenance and embryo implantation, partly due to the synthesis of progesterone. It has been demonstrated that hypoxia regulates invasion, proliferation, and differentiation of trophoblast cells. Additionally, human trophoblasts display rhythmic expression of circadian clock genes. However, it remains unclear if the circadian clock system is present in goat trophoblast cells (GTCs), and its involvement in hypoxia regulation of steroid hormone synthesis remains elusive. In this study, immunofluorescence staining revealed that both BMAL1 and NR1D1 (two circadian clock components) were highly expressed in GTCs. Quantitative real-time PCR analysis showed that several circadian clock genes were rhythmically expressed in forskolin-synchronized GTCs. To mimic hypoxia, GTCs were treated with hypoxia-inducing reagents (CoCl2 or DMOG). Quantitative real-time PCR results demonstrated that hypoxia perturbed the mRNA expression of circadian clock genes and StAR. Notably, the increased expression of NR1D1 and the reduction of StAR expression in hypoxic GTCs were also detected by western blotting. In addition, progesterone secretion exhibited a notable decline in hypoxic GTCs. SR9009, an NR1D1 agonist, significantly decreased StAR expression at both the mRNA and protein levels and markedly inhibited progesterone secretion in GTCs. Moreover, SR8278, an NR1D1 antagonist, partially reversed the inhibitory effect of CoCl2 on mRNA and protein expression levels of StAR and progesterone synthesis in GTCs. Our results demonstrate that hypoxia reduces StAR expression via the activation of NR1D1 signaling in GTCs, thus inhibiting progesterone synthesis. These findings provide new insights into the NR1D1 regulation of progesterone synthesis in GTCs under hypoxic conditions.

Detection of Antibiotic Resistance in Feline-Origin ESBL Escherichia coli from Different Areas of China and the Resistance Elimination of Garlic Oil to Cefquinome on ESBL E. coli.

The development of drug-resistance in the opportunistic pathogen Escherichia coli has become a global public health concern. Due to the share of similar flora between pets and their owners, the detection of pet-origin antibiotic-resistant E. coli is necessary. This study aimed to detect the prevalence of feline-origin ESBL E. coli in China and to explore the resistance elimination effect of garlic oil to cefquinome on ESBL E. coli. Cat fecal samples were collected from animal hospitals. The E. coli isolates were separated and purified by indicator media and polymerase chain reaction (PCR). ESBL genes were detected by PCR and Sanger sequencing. The MICs were determined. The synergistic effect of garlic oil and cefquinome against ESBL E. coli was investigated by checkerboard assays, time-kill and growth curves, drug-resistance curves, PI and NPN staining, and a scanning electronic microscope. A total of 80 E. coli strains were isolated from 101 fecal samples. The rate of ESBL E. coli was 52.5% (42/80). The prevailing ESBL genotypes in China were CTX-M-1, CTX-M-14, and TEM-116. In ESBL E. coli, garlic oil increased the susceptibility to cefquinome with FICIs from 0.2 to 0.7 and enhanced the killing effect of cefquinome with membrane destruction. Resistance to cefquinome decreased with treatment of garlic oil after 15 generations. Our study indicates that ESBL E. coli has been detected in cats kept as pets. The sensitivity of ESBL E. coli to cefquinome was enhanced by garlic oil, indicating that garlic oil may be a potential antibiotic enhancer.

Ophiopogon Polysaccharide Liposome Regulated the Immune Activity of Kupffer Cell through miR-4796.

The purpose of this article is to study the effects and mechanism of miR-4796 in the process of ophiopogon polysaccharide liposome (OPL) regulation of the immune activity of Kupffer cells (KCs). In this study, KCs were used as cell models, and were treated with OPL in different concentrations after being transfected with miR-4796 mimic or miR-4796 inhibitor. Firstly, the secretion of NO and iNOS, phagocytic activity, the expression of surface molecules CD14 and MHC II, apoptosis and ROS secretion were measured by Griess, flow cytometry, fluorescence staining and ELISA. Then, real-time PCR and Western blot were used to measure the expression of TLR4, IKKß, MyD88 and NF-κB in the TLR4-NF-κB signaling pathway. The results showed that after transfection with miR-4796 mimic, the secretion of NO and iNOS, cell migration, cell phagocytosis and expression levels of CD14 and MHC II in the OPL group were significantly higher than those in the miR-4796 mimic control group (p < 0.05; p < 0.01). In addition, the mRNA and protein expression levels of TLR4, MyD88 and NF-κB were significantly higher than those in miR-4796 mimic control group (p < 0.05; p < 0.01). After transfection with miR-4796 inhibitor, the secretion of NO and iNOS, cell migration, cell phagocytosis, expression of CD14 and MHCII in OPL group were significantly higher than those in the miR-4796 inhibitor control group (p < 0.05; p < 0.01). These results indicated that OPL could regulate the immune activity of KCs by regulating miR-4796 and activating the TLR4-NF-κB signaling pathway.

AMP-Activated Protein Kinase Mediates the Antiplatelet Effects of the Thiazolidinediones Rosiglitazone and Pioglitazone.

The thiazolidinedione antidiabetic drugs rosiglitazone and pioglitazone exert antiplatelet effects. Such effects are known to be mediated by the peroxisome proliferator-activated receptor γ (PPARγ), an acknowledged target of the thiazolidinediones, although the molecular mechanism is elusive. Recently, AMP-activated protein kinase (AMPK) signaling was reported to inhibit platelet aggregation. Because AMPK is another target of the thiazolidinediones, the impact of rosiglitazone and pioglitazone on platelet AMPK and its involvement in aggregation were investigated to assess the contribution of AMPK to the antiplatelet activity of these agents. Treatment with rosiglitazone stimulated both AMPK and PPARγ in isolated rat platelets. However, the concentration and the treatment time required for activation were distinct from each other. Indeed, stimulation of AMPK and PPARγ were discrete events without any cross-activation in platelets. Activation of AMPK or PPARγ by rosiglitazone rendered platelets less responsive to aggregatory stimuli such as collagen, ADP, and thrombin. However, the resultant efficacy caused by activating AMPK was higher than that attributable to PPARγ stimulation. Similar results were obtained with pioglitazone. Taken together, rosiglitazone and pioglitazone inhibit platelet aggregation by activating AMPK. AMPK functions as a potential target of rosiglitazone and pioglitazone for their antiplatelet activity, although the in vivo or clinical relevance remains to be assessed.

Reduction of TRAIL-induced Mcl-1 and cIAP2 by c-Myc or sorafenib sensitizes resistant human cancer cells to TRAIL-induced death.

Cells expressing oncogenic c-Myc are sensitized to TNF superfamily proteins. c-Myc also is an important factor in determining whether a cell is sensitive to TRAIL-induced apoptosis, and it is well established that the mitochondrial pathway is essential for apoptosis induced by c-Myc. We investigated whether c-Myc action on the mitochondria is required for TRAIL sensitivity and found that Myc sensitized cells with defective intrinsic signaling to TRAIL. TRAIL induced expression of antiapoptotic Mcl-1 and cIAP2 through activation of NF-kappaB. Both Myc and the multikinase inhibitor sorafenib block NF-kappaB. Combining sorafenib with TRAIL in vivo showed dramatic efficacy in TRAIL-resistant tumor xenografts. We propose the combination of TRAIL with sorafenib holds promise for further development.

Graphenol defects induced blue emission enhancement in chemically reduced graphene quantum dots.

In this work, few layer graphene quantum dots (GQDs) with a size of 3-5 nm are purposely treated with highly concentrated aqueous NaBH4 solutions to obtain the reduced graphene quantum dots (rGQDs). Fourier transform infrared spectroscopy (FT-IR), X-ray photoelectron spectroscopy (XPS) and Raman spectroscopy demonstrate that the number of carbonyl groups decreases but -OH related defects increase during chemical reduction. Green and weak emissions of original GQDs originate from carrier recombination in the disorder-induced localized state (mainly including carbonyl and carboxyl and epoxy groups). As the reduction degree increases, the photoluminescence (PL) quantum efficiency of GQDs increases dramatically from 2.6% to 10.1%. In the meantime, the PL peak position blue shifts rapidly, and full width at half maximum (FWHM) becomes narrower. Thus we can infer that graphenol topological defects (hydroxyl functionalized graphene) are gradually formed during reduction. Besides, graphenol defect related PL features a longer fluorescence lifetime, excitation wavelength dependence but less pH sensitivity.

Mitofusin 2-containing mitochondrial-reticular microdomains direct rapid cardiomyocyte bioenergetic responses via interorganelle Ca(2+) crosstalk.

RATIONALE: Mitochondrial Ca(2+) uptake is essential for the bioenergetic feedback response through stimulation of Krebs cycle dehydrogenases. Close association of mitochondria to the sarcoplasmic reticulum (SR) may explain efficient mitochondrial Ca(2+) uptake despite low Ca(2+) affinity of the mitochondrial Ca(2+) uniporter. However, the existence of such mitochondrial Ca(2+) microdomains and their functional role are presently unresolved. Mitofusin (Mfn) 1 and 2 mediate mitochondrial outer membrane fusion, whereas Mfn2 but not Mfn1 tethers endoplasmic reticulum to mitochondria in noncardiac cells. OBJECTIVE: To elucidate roles for Mfn1 and 2 in SR-mitochondrial tethering, Ca(2+) signaling, and bioenergetic regulation in cardiac myocytes. METHODS AND RESULTS: Fruit fly heart tubes deficient of the Drosophila Mfn ortholog MARF had increased contraction-associated and caffeine-sensitive Ca(2+) release, suggesting a role for Mfn in SR Ca(2+) handling. Whereas cardiac-specific Mfn1 ablation had no effects on murine heart function or Ca(2+) cycling, Mfn2 deficiency decreased cardiomyocyte SR-mitochondrial contact length by 30% and reduced the content of SR-associated proteins in mitochondria-associated membranes. This was associated with decreased mitochondrial Ca(2+) uptake (despite unchanged mitochondrial membrane potential) but increased steady-state and caffeine-induced SR Ca(2+) release. Accordingly, Ca(2+)-induced stimulation of Krebs cycle dehydrogenases during ß-adrenergic stimulation was hampered in Mfn2-KO but not Mfn1-KO myocytes, evidenced by oxidation of the redox states of NAD(P)H/NAD(P)(+) and FADH(2)/FAD. CONCLUSIONS: Physical tethering of SR and mitochondria via Mfn2 is essential for normal interorganelle Ca(2+) signaling in the myocardium, consistent with a requirement for SR-mitochondrial Ca(2+) signaling through microdomains in the cardiomyocyte bioenergetic feedback response to physiological stress.

Preparation, characteristic, biological activities, and application of polysaccharide from Lilii Bulbus: a review.

OBJECTIVES: This review highlights the current knowledge of polysaccharide from Lilii Bulbus, including the extraction, purification, structure, structure modification , biological activities and application, which will hopefully provide reference for further research and development of polysaccharide from Lilii Bulbus. MATERIALS AND METHODS: Literature searches were conducted on the following databases: Pubmed, ACS website, Elsevier, Google Scholar, Web of Science and CNKI database. Keywords such as "Lilii Bulbus", "polysaccharide", "preparation", "biological activities" and "application" were used to search relevant journals and contents, and some irrelevant contents were excluded. RESULTS: In general, the study of Lilium Bulbus polysaccharide extraction and purification, structure characterization and biological activity has made substantial progress, these findings highlight the lilium brownii polysaccharide enormous potential in biomedical applications, of lilium brownii polysaccharide laid a solid foundation for further research. DISCUSSION AND CONCLUSIONS: However, it should be noted that the relevant mechanism of the effective effect of lily bulb polysaccharide still needs to be worked on by researchers. These findings highlight the great potential of lily polysaccharides in biomedical applications, and lay a solid foundation for further research on lily polysaccharides.

Salvia miltiorrhiza ameliorates endometritis in dairy cows by relieving inflammation, energy deficiency and blood stasis.

Introduction: According to traditional Chinese veterinary medicine, endometritis is caused by a combination of Qi deficiency, blood stasis, and external evil invasion. Salvia miltiorrhiza is a traditional Chinese medicine that counteracts blood stasis and has additional demonstrated effects in boosting energy and restraining inflammation. Salvia miltiorrhiza has been employed in many traditional Chinese prescriptions that have proven effective in healing clinical dairy cow endometritis. Methods: the in vivo effect of Salvia miltiorrhiza in treating endometritis was evaluated in dairy cows. In addition, bovine endometrial epithelium cell inflammation and rat blood stasis models were employed to demonstrate the crosstalk between energy, blood circulation and inflammation. Network analysis, western blotting, qRT-PCR and ELISA were performed to investigate the molecular mechanism of Salvia miltiorrhiza in endometritis treatment. Results: The results demonstrate that treatment with Salvia miltiorrhiza relieves uterine inflammation, increases blood ATP concentrations, and prolongs blood clotting times. Four of the six Salvia miltiorrhiza main components (SMMCs) (tanshinone IIA, cryptotanshinone, salvianolic acid A and salvianolic acid B) were effective in reversing decreased ATP and increased IL-1ß, IL-6, and IL-8 levels in an in vitro endometritis model, indicating their abilities to ameliorate the negative energy balance and external evil invasion effects of endometritis. Furthermore, in a blood stasis rat model, inflammatory responses were induced in the absence of external infection; and all six SMMCs inhibited thrombin-induced platelet aggregation. Network analysis of SMMC targets predicted that Salvia miltiorrhiza may mediate anti-inflammation via the Toll-like receptor signaling pathway; anti-aggregation via the Platelet activation pathway; and energy balance via the Thermogenesis and AMPK signaling pathways. Multiple molecular targets within these pathways were verified to be inhibited by SMMCs, including P38/ERK-AP1, a key molecular signal that may mediate the crosstalk between inflammation, energy deficiency and blood stasis. Conclusion: These results provide mechanistic understanding of the therapeutic effect of Salvia miltiorrhiza for endometritis achieved through Qi deficiency, blood stasis, and external evil invasion.

Polygonum cillinerve polysaccharide inhibits transmissible gastroenteritis virus by regulating microRNA-181.

Transmissible gastroenteritis virus (TGEV) is an important pathogen capable of altering the expression profile of cellular miRNA. In this study, the potential of Polygonum cillinerve polysaccharide (PCP) to treat TGEV-infected piglets was evaluated through in vivo experiments. High-throughput sequencing technology was employed to identify 9 up-regulated and 17 down-regulated miRNAs during PCP-mediated inhibition of TGEV infection in PK15 cells. Additionally, miR-181 was found to be associated with target genes of key proteins in the apoptosis pathway. PK15 cells were treated with various concentrations of PCP following transfection with miR-181 mimic or inhibitor. Real-time PCR assessed the impact on TGEV replication, while electron microscopy (TEM) and Hoechst fluorescence staining evaluated cellular functionality. Western blot analysis was utilized to assess the expression of key signaling factors-cytochrome C (cyt C), caspase 9, and P53-in the apoptotic signaling pathway. The results showed that compared with the control group, 250 µg/mL PCP significantly inhibited TGEV gRNA replication and gene N expression (P < 0.01). Microscopic examination revealed uniform cell morphology and fewer floating cells in PCP-treated groups (250 and 125 µg/mL). TEM analysis showed no typical virus structure in the 250 µg/mL PCP group, and apoptosis staining indicated a significant reduction in apoptotic cells at this concentration. Furthermore, PCP may inhibit TGEV-induced apoptosis via the Caspase-dependent mitochondrial pathway following miR-181 transfection. These findings provide a theoretical basis for further exploration into the mechanism of PCP's anti-TGEV properties.

Effect of miR-17 on Polygonum Cillinerve polysaccharide against transmissible gastroenteritis virus.

Transmissible gastroenteritis virus (TGEV) could cause diarrhea, vomiting, dehydration and even death in piglets, miRNA played an important role in the interaction between virus and cell. The study aimed to investigate the impact of miR-17 on the polysaccharide of Polygonum Cillinerve (PCP) in combating TGEV. miR-17 was screened and transfection validation was performed by Real-time PCR. The function of miR-17 on PK15 cells infected with TGEV and treated with PCP was investigated by DCFH-DA loading probe, JC-1 staining and Hoechst fluorescence staining. Furthermore, the effect of miR-17 on PCP inhibiting TGEV replication and apoptosis signaling pathways during PCP against TGEV infection was measured through Real-time PCR and Western blot. The results showed that miR-17 mimic and inhibitor could be transferred into PK15 cells and the expression of miR-17 significantly increased and decreased respectively compared with miR-17 mimic and inhibitor (P < 0.05). A total 250 µg/mL of PCP could inhibit cells apoptosis after transfection with miR-17. PCP (250 µg/mL and 125 µg/mL) significantly inhibited the decrease in mitochondrial membrane potential induced by TGEV after transfection with miR-17 (P < 0.05). After transfection of miR-17 mimic, PCP at concentrations of 250 µg/mL and 125 µg/mL significantly promoted the mRNA expression of P53, cyt C and caspase 9 (P < 0.05). Compared with the control group, the replication of TGEV gRNA and gene N was significantly inhibited by PCP at concentrations of 250 µg/mL and 125 µg/mL after transfection of both miR-17 mimic and inhibitor (P < 0.05). PCP at 62.5 µg/mL significantly inhibited the replication of gene S following transfection with miR-17 inhibitor (P < 0.05). These results suggested that PCP could inhibit the replication of TGEV and apoptosis induced by TGEV by regulating miR-17.

Effect of different acupuncture sequences of Huiyangjiuzhen acupoints on blood glucose and hemorheology in the anesthetized rabbits.

Background and objective: Hemorheology and blood glucose are commonly used to estimate the risks of thrombosis and stress hyperglycemia after anaesthesia. The sequence of acupoint stimulation might influence the therapeutic effects of acupuncture. In the current study, we aimed at investigating the effect of different acupuncture sequences of "Huiyangjiuzhen" acupoints on the blood glucose and hemorheology in anesthetized rabbits. Methods: Twenty-five rabbits were randomly divided into five groups, including the control group (CG), the positive-sequence group (PSG), the reverse-sequence group (RSG), the disorder-sequence group (DSG), and the random group (RG). Except for the CG and RG, the rabbits in other groups were acupunctured with different sequences of "Huiyangjiuzhen"acupoints when the rabbits were anesthetized. The acupoints in rabbits of the RG were chosen randomly. The levels of blood glucose and hemorheology indexes before and after anaesthesia was detected. Results: In the PSG, Hηb 200/s, Mηb 30/s, Hηr 200/s, ERI, hematocrit and plasma viscosity levels were decreased, and the blood glucose level was not changed. In the DSG, the levels of Mηb 30/s and hematocrit were decreased, and the blood glucose was increased. In the CG, RSG and RG, no hemorheology indexes were changed and the blood glucose was increased. Conclusion: "Huiyangjiuzhen" acupuncture could decrease the risks of post-operative thrombosis and stress hyperglycemia in anesthetized rabbits. This effectiveness depends on both acupuncture and acupuncture sequence at the "Huiyangjiuzhen" acupoints.

Mitochondrial fusion is essential for organelle function and cardiac homeostasis.

RATIONALE: Mitochondria constitute 30% of myocardial mass. Mitochondrial fusion and fission appear essential for health of most tissues. Mitochondrial fission occurs in neonatal cardiomycyte and is implicated in cardiomyocyte death. Mitochondrial fusion has not been observed in postmitotic myocytes of adult hearts, and its occurrence and function in this context are controversial. OBJECTIVE: Determine the consequences on organelle and organ function of disrupting cardiomyocyte mitochondrial fusion in vivo. METHODS AND RESULTS: The murine mfn1 and mfn2 genes, encoding mitofusins (Mfn) 1 and 2 that mediate mitochondrial tethering and outer mitochondrial membrane fusion, were interrupted by Cre-mediated excision of essential exons in neonatal (Nkx2.5-Cre) and adult (MYH6 modified estrogen receptor-Cre-modified estrogen receptor plus tamoxifen or Raloxifene) hearts. Embryonic combined Mfn1/Mfn2 ablation was lethal after e9.5. Conditional combined Mfn1/Mfn2 ablation in adult hearts induced mitochondrial fragmentation, cardiomyocyte and mitochondrial respiratory dysfunction, and rapidly progressive and lethal dilated cardiomyopathy. Before heart failure developed, cardiomyocyte shortening and calcium cycling were unaffected by absence of Mfn1 and Mfn2. Based on the time course over which fusion-defective mitochondrial size decreases, a mitochondrial fusion/fission cycle in adult mouse hearts occurs approximately every 16 days. CONCLUSIONS: Mitochondrial fusion in adult cardiac myocytes is necessary to maintain normal mitochondrial morphology and is essential for normal cardiac respiratory and contractile function. Interruption of mitochondrial fusion causes lethal cardiac failure at a time corresponding to 3 or 4 cycles of unopposed mitochondrial fission.

Research progress on the extraction technology and activity study of Epimedium polysaccharides.

More pharmacological effects of polysaccharides from traditional Chinese medicines have been discovered in recent years. Epimedium has been used for thousands of years as a traditional Chinese medicine in China. Water-soluble Epimedium polysaccharides is one of the main ingredients of Epimedium, which is one of the main active ingredients of Epimedium, mainly composed of mannose, rhamnose, galacturonic acid, glucose, and galactose. The extraction methods of Epimedium polysaccharides including hot water extraction, cellulase extraction, ultrasonic extraction, microwave-assisted extraction, ultrasound compound enzyme and ultra-high pressure extraction, they affect the yield of Epimedium polysaccharides. The characteristics of deproteinization including enzyme deproteinization, macroporous resin deproteinization and Sevag methods are introduced respectively. Some chemical modification methods of Epimedium polysaccharides are also involved such as phosphorylation, sulfation, selenization, and lipids encapsulated. Epimedium polysaccharides have a variety of pharmacological activities, including immune promotion, reproduction promotion, anti-osteoporosis, anti-tumor, antioxidant, anti-fatigue and antivirus, also beneficial to nervous and hematopoietic systems. At present, the research of Epimedium polysaccharides has been in depth. In this paper, the research progress on extraction, purification, chemical modification methods and pharmacological activity of Epimedium polysaccharides summarized. The aim is to provide reference for further research and development of Epimedium polysaccharides.

Four Patterns of Canine Wei Syndrome Treated with Traditional Chinese Medicine.

Atrophy and weakness of the limbs is a common condition in animals, especially dogs. It typically presents with flaccidity and weakness of the limbs, especially the hind legs, muscle atrophy, and the inability to walk. In Traditional Chinese Medicine (TCM) and Traditional Chinese Veterinary Medicine (TCVM), this is known as wei syndrome (WS). According to TCM, the etiology of WS can be (1) lung heat and fluid consumption; (2) insufficiency of the liver and kidneys; (3) dampness-heat invasion; (4) damage to the spleen and stomach, which are also the patterns of WS. This report aims to provide an alternative option for the treatment of canine paralysis. Four dogs with different WS patterns were treated with acupuncture, moxibustion, and Chinese herbs based on the guidelines of the TCM literature. Three patients recovered normal functioning. The fourth patient could walk normally after 2 weeks of treatment, but his hind limbs became weak again 3 months later. Weekly acupuncture treatment was resumed until his death 18 months later. TCM application of acupuncture, moxibustion, and Chinese herbs can be an effective treatment for canine WS. It is hoped that this case report will broaden the treatment options of other veterinarians when patients present with this condition.

Genome characteristics of the optrA-positive Clostridium perfringens strain QHY-2 carrying a novel plasmid type.

Clostridium perfringens is a bacterial species of importance to both public and animal health. The gene optrA is the first gene that confers resistance to the tedizolid, a last-resort antimicrobial agent in human medicine. Herein, we whole-genome sequenced and analyzed one optrA-positive C. perfringens strain QHY-2 from Tibetan sheep in Qinghai province and identified one optrA plasmid pQHY-2. The plasmid shared similar structure with the optrA-positive plasmids p2C45 and p21-D-5b previously identified in C. perfringens, demonstrating the potential horizontal transmission of the optrA plasmids among C. perfringens strains. Annotation of the optrA-positive plasmids showed optrA and erm(A) located on a segment flanked by IS element IS1216E, and fexA, optrA, and erm(A) located on a segment flanked by IS element ISVlu1, which revealed the possible dissemination mechanism. Additionally, a Tn6218-like transposon carrying aac(6')-aph(2″) and erm(B) was also detected on pQHY-2, demonstrating the transposition of Tn6218 and spread of antibiotic resistance among Clostridium bacteria. Molecular analysis indicated the optrA-positive plasmids belonged to a plasmid type distinct from the pCW3-like plasmids, pCP13-like plasmids, or pIP404-like plasmids. Further structure analysis showed they might be formed by inserting segments into plasmid pCPCPI53k-r1_1, which coexist with two pCW3-like plasmids and one pCP13-like plasmid in C. perfringens strain CPI 53k-r1 isolated from a healthy human in Finland. IMPORTANCE Antimicrobial resistance is now a global concern posing threats to food safety and public health. The pCW3-like plasmids can encode several main toxin genes and three antibiotic resistance genes (ARGs), including tetA(P), tetB(P), and erm(B), which used to be considered as the main carrier of ARGs in Clostridium perfringens. In this study, we found the optrA plasmids, which belonged to a novel plasmid type, could also harbor many other ARGs, indicating this type of plasmid might be the potential repository of ARGs in C. perfringens. Additionally, this type of plasmid could coexist with the pCW3-like plasmids and pCP13-like plasmids that encoded toxin genes associated with gastrointestinal diseases, which showed the potential threat to public health.

Detection of antibiotic-resistant canine origin Escherichia coli and the synergistic effect of magnolol in reducing the resistance of multidrug-resistant Escherichia coli.

Background: The development of antimicrobial resistance in the opportunistic pathogen Escherichia coli has become a global public health concern. Due to daily close contact, dogs kept as pets share the same E. coli with their owners. Therefore, the detection of antimicrobial resistance in canine E. coli is important, as the results could provide guidance for the future use of antibiotics. This study aimed to detect the prevalence of antibiotic-resistance of canine origin E. coli in Shaanxi province and to explore the inhibition effect of magnolol combined with cefquinome on MDR E. coli, so as to provide evidence for the use of antibiotics. Methods: Canine fecal samples were collected from animal hospitals. The E. coli isolates were separated and purified using various indicator media and polymerase chain reaction (PCR). Drug-resistance genes [aacC2, ant(3')-I, aph(3')-II, aac(6')-Ib-cr, aac(3')-IIe, bla KPC , bla IMP-4 , bla OXA , bla CMY , bla TEM-1 , bla SHV , bla CTX-M-1 , bla CTX-M-9 , Qnra, Qnrb, Qnrs, TetA, TetB, TetM, Ermb] were also detected by PCR. The minimum inhibitory concentration (MIC) was determined for 10 antibiotics using the broth-microdilution method. Synergistic activity of magnolol and cefquinome against multidrug-resistant (MDR) E. coli strains was investigated using checkerboard assays, time-kill curves, and drug-resistance curves. Results: A total of 101 E. coli strains were isolated from 158 fecal samples collected from animal hospitals. MIC determinations showed that 75.25% (76/101) of the E. coli strains were MDR. A total of 22 drug-resistance genes were detected among the 101 strains. The bla TEM-1gene exhibited the highest detection rate (89.77%). The TetA and Sul gene also exhibited high detection rate (66.34 and 53.47%, respectively). Carbapenem-resistant E. coli strains were found in Shangluo and Yan'an. Additionally, in MDR E. coli initially resistant to cefquinome, magnolol increased the susceptibility to cefquinome, with an FICI (Fractional Inhibitory Concentration Index) between 0.125 and 0.5, indicating stable synergy. Furthermore, magnolol enhanced the killing effect of cefquinome against MDR E. coli. Resistance of MDR E. coli to cefquinome decreased markedly after treatment with magnolol for 15 generations. Conclusion: Our study indicates that antibiotic-resistance E. coli has been found in domestic dogs. After treatment with magnolol extracted from the Chinese herb Houpo (Magnolia officinalis), the sensitivity of MDR E. coli to cefquinome was enhanced, indicating that magnolol reverses the resistance of MDR E. coli. The results of this study thus provide reference for the control of E. coli resistance.

Extraction, purification, structural character and biological properties of propolis flavonoids: A review.

Propolis is an aromatic substance which is collected by bees and mixed with bee saliva. The plant sources of propolis are mainly consisted with plant exudates from bark, buds and etc. Flavonoids are secondary metabolites widely found in natural plants, which have a variety of health care functions and are the main active ingredients of propolis. This article summarized the types, active ingredients, pharmacological effects, extraction methods and applications of propolis flavonoids, the aim was to provide the theoretical basis for further research and development of propolis flavonoids.

Study on the SHP2-Mediated Mechanism of Promoting Spermatogenesis Induced by Active Compounds of Eucommiae Folium in Mice.

Spermatogenesis directly determines the reproductive capacity of male animals. With the development of society, the increasing pressure on people's lives and changes in the living environment, male fertility is declining. The leaf of Eucommia ulmoides Oliv. (Eucommiae Folium, EF) was recorded in the 2020 Chinese Pharmacopoeia and was used in traditional Chinese medicine as a tonic. In recent years, EF has been reported to improve spermatogenesis, but the mechanisms of EF remain was poorly characterized. In this study, the effect of EF ethanol extract (EFEE) on spermatogenesis was tested in mice. Chemical components related to spermatogenesis in EF were predicted by network pharmacology. The biological activity of the predicted chemical components was measured by the proliferation of C18-4 spermatogonial stem cells (SSCs) and the testosterone secretion of TM3 leydig cells. The biological activity of chlorogenic acid (CGA), the active compound in EF, was tested in vivo. The cell cycle was analysed by flow cytometry. Testosterone secretion was detected by ELISA. RNA interference (RNAi) was used to detect the effect of key genes on cell biological activity. Western blotting, qRT-PCR and immunofluorescence staining were used to analyse the molecular mechanism of related biological activities. The results showed that EFEE and CGA could improve spermatogenesis in mice. Furthermore, the main mechanism was that CGA promoted SSC proliferation, self-renewal and Leydig cell testosterone secretion by promoting the expression of SHP2 and activating the downstream signaling pathways involved in these biological processes. This study provided strong evidence for elucidating the mechanism by which EF promotes the spermatogenesis in mice and a new theoretical basis for dealing with the decrease in male reproductive capacity.