In order to establish a simple,sensitive and specific diagnostic method for the simultane-ous detection of Mycoplasma hyorhinis(Mhr)and Mycoplasma hyopneumoniae(Mhp)in swine,specific primers and probes were designed using the Mhr p37 and Mhp p36 gene sequences as the target genes,and the dual LFD-RPA rapid test was established by screening the primers and probes,optimizing primer ratios and evaluating its effectiveness through the sensitivity,reproduc-ibility and clinical sample testing.The sensitivity,specificity,reproducibility and clinical samples were evaluated.The results showed that the established dual LFD-RPA assay could complete the amplification in 15 min at 39 ℃,and its optimal primer ratio was 1.6∶0.8,and the lowest detection limits were up to 3.63 and 3.60 copies/μL,respectively;the reproducibility was stable;and there was no cross-reactivity with Pasteurella multocida,Bordetella bronchiseptica,Haemophilus pa-rasuis,Actinobacillus pleuropneumoniae,Escherichia coli.The test successfully established a dual LFD-RPA assay,which can detect Mhr and Mhp simultaneously,and is simple,sensitive and spe-cific without relying on specialized equipment,and is suitable for carrying out on-site rapid diagno-sis of Mhr and Mhp.

In order to establish a simple,sensitive and specific diagnostic method for the simultane-ous detection of Mycoplasma hyorhinis(Mhr)and Mycoplasma hyopneumoniae(Mhp)in swine,specific primers and probes were designed using the Mhr p37 and Mhp p36 gene sequences as the target genes,and the dual LFD-RPA rapid test was established by screening the primers and probes,optimizing primer ratios and evaluating its effectiveness through the sensitivity,reproduc-ibility and clinical sample testing.The sensitivity,specificity,reproducibility and clinical samples were evaluated.The results showed that the established dual LFD-RPA assay could complete the amplification in 15 min at 39 ℃,and its optimal primer ratio was 1.6∶0.8,and the lowest detection limits were up to 3.63 and 3.60 copies/μL,respectively;the reproducibility was stable;and there was no cross-reactivity with Pasteurella multocida,Bordetella bronchiseptica,Haemophilus pa-rasuis,Actinobacillus pleuropneumoniae,Escherichia coli.The test successfully established a dual LFD-RPA assay,which can detect Mhr and Mhp simultaneously,and is simple,sensitive and spe-cific without relying on specialized equipment,and is suitable for carrying out on-site rapid diagno-sis of Mhr and Mhp.

Medical institutions, with their clinical practice foundation and abundant human use experience data, have become important carriers for the inheritance and innovation of traditional Chinese medicine(TCM) and the "cradles" of the preparation of new TCM. To effectively promote the transformation of new TCM originating from the TCM clinical practice in medical institutions and establish an effective evaluation index system for the transformation of new TCM conforming to the characteristics of TCM, consensus experts adopted the literature research, questionnaire survey, Delphi method, etc. By focusing on the policy and technical evaluation of new TCM originating from the TCM clinical practice in medical institutions, a comprehensive evaluation from the dimensions of drug safety, efficacy, feasibility, and characteristic advantages was conducted, thus forming a comprehensive evaluation system with four primary indicators and 37 secondary indicators. The expert consensus reached aims to encourage medical institutions at all levels to continuously improve the high-quality research and development and transformation of new TCM originating from the TCM clinical practice in medical institutions and targeted at clinical needs, so as to provide a decision-making basis for the preparation, selection, cultivation, and transformation of new TCM for medical institutions, improve the development efficiency of new TCM, and precisely respond to the public medication needs.
Medicine, Chinese Traditional/standards* ; Humans ; Consensus ; Drugs, Chinese Herbal/therapeutic use* ; Surveys and Questionnaires

Notum, a negative feedback regulator of the Wnt signaling, has emerged as a promising target for treating glucocorticoid-induced osteoporosis (GIOP). This study showcases an efficient strategy for discovering the anti-Notum constituents from herbal medicines (HMs) as novel anti-GIOP agents. Firstly, a rapid-responding near-infrared fluorogenic substrate for Notum was rationally engineered for high-throughput identifying the anti-Notum HMs. The results showed that Bu-Gu-Zhi (BGZ), a known anti-osteoporosis herb, potently inhibited Notum in a competitive-inhibition manner. To uncover the key anti-Notum constituents in BGZ, an efficient strategy was adapted via integrating biochemical, phytochemical, computational, and pharmacological assays. Among all identified BGZ constituents, three furanocoumarins were validated as strong Notum inhibitors, while 5-methoxypsoralen (5-MP) showed the most potent anti-Notum activity and favorable safety profiles. Mechanistically, 5-MP acted as a competitive inhibitor of Notum via creating strong hydrophobic interactions with Trp128 and Phe268 in the catalytic cavity of Notum. Cellular assays showed that 5-MP remarkably promoted osteoblast differentiation and activated Wnt signaling in dexamethasone (DXMS)-challenged MC3T3-E1 osteoblasts. In dexamethasone-induced osteoporotic mice, 5-MP strongly elevated bone mineral density (BMD) and improved cancellous and cortical bone thickness. Collectively, this study constructs a high-efficient platform for discovering key anti-Notum constituents from HMs, while 5-MP emerges as a promising anti-GIOP agent.

Objective To investigate the expression pattern of GFAP and VEGF after closed brain injury in rats,and to infer the time elapsed after brain injury,and then to provide reference for the inference of injury time.Methods Male rats were randomly divided into control group and experimental group,in which the experimental group was divided into 1 h,6 h,12 h,1 d,3 d,7 d and 14 d groups according to the different elapsed time after injury.Immunohistochemistry and ELISA were used to detect the expression of GFAP and VEGF in brain tissue and serum of rats after closed brain injury.Results(1)In each immunohistochemistry experimental group,GFAP was mainly expressed in the cytoplasm of astrocytes,with an obvious increase at 1 h after injury,a progressive increase of expression in 7 d,and a decrease of expression close to the level of the control group at 14 d after injury.While VEGF was mainly expressed in the cytoplasm of neurons,with an obvious increase at 1 d after injury,a peak at 3 d post-injury,and a decrease in expression at 14 d.(2)In each ELISA experimental group,the VEGF content in serum of each experimental group increased earlier than the VEGF expression in brain tissue,which began to increase significantly at 6 h after injury,and the content was the most at 3 d after injury,and was close to that of the control group at 14 d after injury.Conclusion(1)It has showed temporal changes in the expression of GFAP and VEGF,and both are expected to be biological markers for inferring the time of injury in forensic work.(2)GFAP and VEGF have high sensitivity and specificity and are important for the extrapolation of early craniocerebral injury time.

Colorectal cancer is a common malignant tumor in the digestive system， ranking third in incidence and second in the cause of death worldwide. In recent years， the incidence of colorectal cancer is on the rise， and the age of patients with colorectal cancer tends to be younger， with a heavy cancer burden. It is of great significance to prevent the occurrence， development， recurrence， and metastasis of colorectal cancer to reduce the incidence and mortality of colorectal cancer. Patriniae Herba has the effects of clearing heat， removing toxins， eliminating carbuncle， and discharging pus and shows good therapeutic efficacy on inflammatory bowel disease， digestive tract tumors， pelvic inflammation， gynecological tumor， and so on. Patriniae Herba is often used in the clinical treatment of colorectal cancer， but its mechanism of action is not clear. Modern studies have found that Patriniae Herba contains triterpenoids， saponins， iridoids， flavonoids， and other chemical components， with antioxidant， anti-tumor， anti-bacterial， and other pharmacological effects. The main anti-tumor components of Patriniae Herba are flavonoids. The analysis of network pharmacology and the spectrum-effect relationship has suggested that quercetin， luteolin， apigenin， isoorientin， and isovitexin play a major role in inhibiting the occurrence and development of colorectal cancer. In vivo and in vitro studies have shown that flavonoids in Patriniae Herba can play an anti-tumor role in various ways， such as preventing precancerous lesions of colorectal cancer， inhibiting the growth and proliferation of cancer cells， blocking cancer cell cycle， promoting cancer cell apoptosis， and reversing drug resistance of colorectal cancer. The oral availability of flavonoids is low. The gut is the main metabolic site of flavonoids in the body， its metabolic pathway is closely related to gut microbiota. This paper reviewed the anti-tumor mechanism of flavonoids and their influence on gut microbiota to provide a reference for further research on the mechanism of Patriniae Herba against colorectal cancer and its clinical application.

Objective To investigate the expression pattern of GFAP and VEGF after closed brain injury in rats,and to infer the time elapsed after brain injury,and then to provide reference for the inference of injury time.Methods Male rats were randomly divided into control group and experimental group,in which the experimental group was divided into 1 h,6 h,12 h,1 d,3 d,7 d and 14 d groups according to the different elapsed time after injury.Immunohistochemistry and ELISA were used to detect the expression of GFAP and VEGF in brain tissue and serum of rats after closed brain injury.Results(1)In each immunohistochemistry experimental group,GFAP was mainly expressed in the cytoplasm of astrocytes,with an obvious increase at 1 h after injury,a progressive increase of expression in 7 d,and a decrease of expression close to the level of the control group at 14 d after injury.While VEGF was mainly expressed in the cytoplasm of neurons,with an obvious increase at 1 d after injury,a peak at 3 d post-injury,and a decrease in expression at 14 d.(2)In each ELISA experimental group,the VEGF content in serum of each experimental group increased earlier than the VEGF expression in brain tissue,which began to increase significantly at 6 h after injury,and the content was the most at 3 d after injury,and was close to that of the control group at 14 d after injury.Conclusion(1)It has showed temporal changes in the expression of GFAP and VEGF,and both are expected to be biological markers for inferring the time of injury in forensic work.(2)GFAP and VEGF have high sensitivity and specificity and are important for the extrapolation of early craniocerebral injury time.

OBJECTIVE: To explore the effectiveness of manipulation treatment for lumbar disc herniation (LDH) based on the model of muscle and bone assessment. METHODS: From May 2022 to August 2023, using the methods single-center randomized controlled in Ruijin Hospital Affiliated to Shanghai Jiaotong University School of Medicine, 72 patients were treated with LDH and divided into muscle and bone assessment model manipulation group and the two step seven gimmick group according to the random number table method, the muscle and bone assessment model manipulation group fall off in 1 case, the two step seven gimmick group falls off in 2 cases. There were 35 cases in the muscle and bone assessment model manipulation group, including 12 males and 23 females;The age was 27 to 48 years old with an average of (37.77±7.63) years old. The course of disease was 35 to 180 days with an average of (83.68±69.01) days. The patients were treated with manual therapy under the guidance of muscle and bone assessment model, twice a week for 4 weeks. There were 34 cases in the two step seven gimmick group including 12 males and 22 females;The age was 26 to 49 years old with an average of (37.59±7.43) years old;The course of disease was 40 to 175 days with an average of (82.15±68.87) days. The patients were treated with two step seven gimmick method, 2 times a week, for 4 weeks. The visual analogue scale (VAS) and Oswestry disability index (Oswestry disability index, ODI) questionnaire, muscle tension and lumbar spine angle and the straight leg-raising activities were compared between two groups before and 4 weeks after treatment. RESULTS: The VAS of the muscle and bone assessment model manipulation group and the two step seven gimmick group(6.51±0.61) and (6.62±0.56) before treatment decreased to 2.40±0.81 and 3.18±0.78 after 4 weeks of treatment, respectively, and the muscle and bone assessment model manipulation group was significantly lower than the two step seven gimmick group (P<0.01). The ODI of the muscle and bone assessment model manipulation group and the two step seven gimmick group were (64.57±5.11) and (65.02±5.18) before treatment, decreased to (18.60±2.27) and (24.70±2.14) after 4 weeks of treatment, and the ODI of the muscle and bone assessment model manipulation group was significantly lower than that of the two step seven gimmick group (P<0.01). Before the treatment, side erector spinae, gluteus medius, and gastrocnemius muscle tension were (59.95±2.60), (62.59±2.51), (49.97±2.01) in the muscle and bone assessment model manipulation group and (60.39±3.84), (62.47±3.27), (49.55±1.27) in the two step seven gimmick group;After 4 weeks of treatment, the muscle tension of erector spinae, gluteus medius and gastrocnemius on the affected side were (56.58±2.71), (60.44±2.31) and (49.19±1.57) in the muscle and bone assessment model manipulation group, (58.28±3.79), (60.11±2.87), (48.55±0.90) in the two step seven gimmick group, the differences had statistical significance before and after treatment of two groups(P<0.01). The muscle and bone assessment model manipulation group was better than the two step seven gimmick group in improving the erector spinae muscle tension on the affected side (P<0.05), and there was no significant difference in the rest (P>0.05). Before the treatment, lumbar proneness, stretch, subject to lateral flexion and lateral angle of the straight leg-raising on the affected side were (46.00±8.89)°, (13.57±3.75)°, (12.29±3.50) °, (43.71±7.98) ° in the muscle and bone assessment model manipulation group, (45.14±6.24) °, (12.23±3.75) °, (12.66±2.98) ° and (44.18±3.50) ° in the two step seven gimmick group. After 4 weeks of treatment, the angles of lumbar flexion, extension, flexion on the affected side and straight leg raising on the affected side were (76.29±4.43) °, (20.00±1.71) °, (22.43±2.81) °, (70.41±7.59) ° in the muscle and bone assessment model manipulation group, and (75.75±6.38) °, (16.43±3.36) °, (20.19±3.52) °, (65.42±6.15) ° in the two step seven gimmick group. The difference had statistical significance before and after treatment in two groups(P<0.01), a comparison between groups, after 4 weeks of treatment, the angles of lumbar flexion and extension, affected side flexion, and lower limb straight leg elevation in the muscle and bone assessment model manipulation group were better than those in the two step seven gimmick group (P<0.05). Before the treatment, pelvic tilt, lumbar lordosis angle were (2.71±1.01) mm, (37.63±3.35) ° in the muscle and bone assessment model manipulation group, and (2.69±0.97) mm, (36.98±3.73) ° in the two step seven gimmick group;After 4 weeks of treatment, the pelvic tilt and lumbar lordosis angle were (0.84±0.36) mm and (41.64±2.96) ° in the muscle and bone assessment model manipulation group, and those in the method of two step seven gimmick group were (1.18±0.75) mm and (41.70±3.14) °. There were significant differences before and after treatment in both groups (P<0.01), and the improvement of pelvic tilt in the muscle and bone assessment model manipulation group was better than that in the method of two step seven gimmick group after 4 weeks of treatment (P<0.05). CONCLUSION The manipulation under the guidance of the muscle and bone assessment model can effectively improve the pain and dysfunction of LDH patients, and has a better effect than the two-step seven-method manipulation group in improving the muscle tension, lumbar motion function and posture.
Humans ; Male ; Female ; Intervertebral Disc Displacement/physiopathology* ; Middle Aged ; Adult ; Lumbar Vertebrae

ObjectiveTo investigate the mechanism of Chaihu Shugansan in the treatment of functional dyspepsia in rats based on mitophagy and PTEN-induced kinase 1 （Pink1）/E3 ubiquitin ligase （Parkin）. signaling pathway. MethodAccording to the principle of random grouping， 40 SD rats were assigned into a normal group， a model group， a Chaihu Shugansan group， and a positive drug （domperidone） group， with 10 rats in each group. The rats in other groups except the normal group received tail-clamping stimulation to replicate the model of functional dyspepsia. After each time of stimulation， the rats in the normal， model， Chaihu Shugansan， and positive drug groups were administrated with normal saline， normal saline， Chaihu Shugansan （4.8 g·kg^-1）， and an aqueous solution of domperidone （4.5 mg·kg^-1）， respectively. After 28 days of modeling， the gastric emptying rate and the small intestine propulsion rate of the rats in each group were measured and the tissue samples were collected. Hematoxylin-eosin （HE） staining was employed for observation of damage in gastric antrum tissue， and transmission electron microscopy （TEM） for ultrastructural observation of gastric interstitial cells of Cajal （ICCs）. Immunofluorescence co-localization was adopted to observe the expression of cytochrome c oxidase （COX Ⅳ） and Parkin. Western blot was employed to determine the expression levels of microtubule-associated protein 1， light chain 3 （LC3）， and the mitophagy-associated proteins prohibitin2 （PHB2）， Pink1， Parkin， and ubiquitin-specific protease 30 （USP30）. ResultCompared with the normal group， the modeling decreased the gastric emptying rate and the small intestine propulsion rate （P<0.05）. Compared with the model group， Chaihu Shugansan increased the gastric emptying rate and the small intestine propulsion rate （P<0.05）. The results of TEM showed that Chaihu Shugansan reduced the swelling degree of mitochondria in gastric antrum tissue. Compared with the normal group， the modeling increased the fluorescence intensity of Parkin in mitochondria （P<0.01）， while such increase can be alleviated by Chaihu Shugansan （P<0.01）. Western blotting results showed that compared with the normal group， the modeling up-regulated the protein levels of LC3， Pink1， Parkin， and PHB2 （P<0.05， P<0.01） and down-regulated the protein level of USP30 （P<0.01）. Compared with the model group， Chaihu Shugansan down-regulated the protein levels of LC3， Pink1， Parkin， and PHB2 （P<0.05， P<0.01） and up-regulated the protein level of USP30 （P<0.01）. ConclusionChaihu Shugansan may treat functional dyspepsia by blocking the Pink1/Parkin signaling pathway to inhibit excessive mitochondrial autophagy in ICCs.

ObjectiveTo study the possible mechanism of Chaihu Shugan Powder （柴胡疏肝散， CSP） in the treatment of functional dyspepsia （FD）. MethodsTwenty-four SD rats were randomly divided into a normal group， a model group， a CSP group and a probiotic group， with six rats in each group.The tail-clamping provocation method was used in all groups except for the normal group to replicate the FD rat model. Simultaneously， the normal group and the model group were given 10 ml/（kg·d） of saline by gavage， while the CSP group and the probiotic group were given 9.6 g/（kg·d） of CSP aqueous decoction and 0.945 g/（kg·d） of probiotic aqueous solution by gavage， respectively， twice daily for four weeks. After four weeks， the gastric emptying and small intestinal propulsion rates were detected in each group of rats. Hematoxylin-eosin （HE） staining was used to observe the histopathological changes in the gastric sinusoids and duodenum of the rats. The changes in the intestinal flora were analyzed by 16s rDNA high-throughput gene sequencing， and the expressions of the duodenal zona occludin 1 （ZO-1） and Occludin were detected by immunohistochemistry and western blotting. Pearson correlation analysis was performed on intestinal flora and ZO-1 and Occludin protein expression. ResultsThe gastric antrum tissue structure was clear in all groups， and the gland structure was regular， with smooth gastric tissue mucosa and no pathological changes such as erosion and ulcer. Compared to those in the normal group， the intestinal villi in the duodenal tissue in the model group were significantly reduced or atrophied， and the goblet cells were arranged in disorder， with eosinophilic infiltration； the gastric emptying rate and small intestinal propulsion rate， as well as ZO-1 and Occludin protein expression in duodenal tissue significantly decreased （P＜0.01）. Compared to those in the model group， the duodenal tissue structure was clear， and the length intestinal villi was longer， with goblet cells neatly arranged in the CSP group and the probiotic group； no obvious eosinophil infiltration was found， and the gastric emptying rate and small intestinal propulsion rate as well as ZO-1 and Occludin protein expression significantly increased in the CSP group； a small amount of eosinophil infiltration was found， and the gastric emptying rate and Occludin protein expression significantly increased in the probiotic group （P＜0.05 or P＜0.01）. Beta diversity analysis of intestinal flora showed that the overall structure of intestinal flora in the model group changed significantly compared to that in the normal group （P＜0.01）. The overall structure of the intestinal flora in the CSP group and the probiotic group was closer to the normal group than the model group. Species composition analysis showed that the relative abundance of the Firmicutes decreased， while the relative abundance of the Bacteroidetes and norank_f_Muribaculaceae increased， and the Bacteroidetes/Firmicutes value increased in the model group than those in the normal group （P＜0.05 or P＜0.01）. Compared to those in the model group， the relative abundance of the Firmicutes increased， while the relative abundance of the Bacteroidetes and norank_f_Muribaculaceae， as well as the Bacteroidetes/Firmicutes value decreased in the CSP group and the probiotic group （P＜0.05 or P＜0.01）. There was no statistically significant difference in each indicator between the probiotic group and the CSP group （P＞0.05）. Pearson correlation analysis showed that at the phylum level， Firmicutes was positively correlated with ZO-1 （r=0.610， P=0.016） and Occludin （r=0.694， P=0.004） protein expression. Bacteroidetes was negatively correlated with ZO-1 （r=-0.557， P=0.031） and Occludin （r=-0.662， P=0.007） protein expression. At the genus level， norank_f_Muribaculaceae was negatively correlated with ZO-1 （r=-0.727， P=0.002） and Occludin （r=-0.760，P=0.001） protein expression. ConclusionCSP can restore the structure of intestinal flora， regulate the abundance levels of Firmicutes， Bacteroidetes and norank_f_Muribaculaceae， up-regulate ZO-1 and Occludin proteins， and thus repairing the duodenal mucosal barrier， and playing a therapeutic role in FD rats.