ObjectiveTo explore the mechanism of Yinchenhao Tang regulating the tumor necrosis factor receptor superfamily member 6 （Fas）/cysteine protease-8 （Caspase-8）/cysteine protease-3 （Caspase-3） signaling pathway to inhibit hepatocyte apoptosis and improve cholestatic liver injury （CLI）. MethodsAmong 48 Wistar rats，12 rats were randomly selected as the blank group，and the other rats were administered alpha-naphthalene isothiocyanate （ANIT） by gavage to induce a CLI model. The modeling rats were randomly divided into the model group， the ursodeoxycholic acid group（0.1 g·kg^-1） and the Yinchenhao Tang group（9.23 g·kg^-1），with 12 rats in each group. The rats in each group were given corresponding drugs by gavage for three consecutive days. The levels of alanine aminotransferase （ALT），aspartate aminotransferase （AST），alkaline phosphatase （ALP），gamma-glutamyl transpeptidase （γ-GT），total bilirubin （TBil） and total bile acid （TBA） in serum were detected. The levels of tumor necrosis factor-α （TNF-α） and interleukin-1β （IL-1β） in liver tissue were detected. The histopathological changes of the liver were observed by hematoxylin-eosin （HE） staining. The protein and mRNA expressions of Fas，Caspase-8，Caspase-3，B-cell lymphoma-2 （Bcl-2） associated X protein （Bax） and Bcl-2 in liver tissue were detected by Western blot and real-time fluorescence quantitative polymerase chain reaction （Real-time PCR）. ResultsCompared with those in the blank group，the levels of ALT，AST，ALP，γ-GT，TBA and TBil in serum of the model group were significantly increased （P<0.01）. The levels and mRNA expressions of TNF-α and IL-1β in liver tissue were significantly increased （P<0.01）. The arrangement of hepatocytes was disordered，and inflammatory cell infiltration and bile duct epithelial cell proliferation were observed. The protein and mRNA expressions of Fas，Caspase-8，Caspase-3 and Bax in liver tissue were significantly increased（P<0.05，P<0.01），while the protein and mRNA expressions of Bcl-2 were significantly decreased （P<0.05，P<0.01）. Compared with those in the model group，the levels of ALP，γ-GT，TBA and TBil in the serum of rats in the ursodeoxycholic acid group were significantly decreased. The levels and mRNA expressions of TNF-α and IL-1β in liver tissue were significantly decreased（P<0.05，P<0.01）. The protein and mRNA expressions of Fas，Caspase-8，Caspase-3 and Bax in liver tissue were significantly decreased （P<0.05，P<0.01），while the mRNA expression of Bcl-2 was significantly increased （P<0.05，）. The levels of ALT，AST，γ-GT，TBA and TBil in the serum of rats in the Yinchenhao Tang group were significantly decreased （P<0.01）. The levels and mRNA expressions of TNF-α and IL-1β in liver tissue were significantly decreased （P<0.05，P<0.01）. The protein expression of Fas and Bax and the mRNA expression of Fas，Caspase-8，Caspase-3 and Bax in liver tissue were significantly decreased （P<0.05，P<0.01），while the protein and mRNA expression of Bcl-2 were significantly increased （P<0.05，P<0.01）. Hepatocyte injury，inflammatory cell infiltration and proliferation of bile duct epithelial cells were reduced. ConclusionYinchenhao Tang can ameliorate CLI，and its mechanism may be related to inhibiting hepatocyte apoptosis mediated by the Fas/Caspase-8/Caspase-3 signaling pathway.

ObjectiveTo study the mechanism of Yinchenhao Tang on the improvement of cholestatic liver injury （CLI） by inhibiting toll-like receptor 4 （TLR4）/myeloid differentiation factor 88 （MyD88）/nuclear transcription factor-κB （NF-κB） pathway via regulating farnesol X receptor （FXR）. MethodsA total of 40 Wistar male rats were randomly selected， with 10 as a blank group，and the remaining rats were subjected to the CLI model induced by alpha-naphthalene isothiocyanate （ANIT）. After modeling，they were randomly divided into the model group， the ursodeoxycholic acid （0.1 g·kg^-1） group and the Yinchenhao Tang （9.23 g·kg^-1） group，with 10 animals in each group. Each administration group was given the corresponding drug by intragastric administration for three consecutive days. Alanine aminotransferase （ALT），aspartate aminotransferase （AST），alkaline phosphatase （ALP），γ-glutamyl transpeptidase （γ-GT），total bile acid （TBA），total bilirubin （TBil） and direct bilirubin （DBil） levels in serum were detected. Tumor necrosis factor-α （TNF-α），interleukin-1β （IL-1β），and interleukin-6 （IL-6） levels in liver tissue were detected. Real-time PCR was used to detect the mRNA expression of FXR，TLR4，MyD88，NF-κB，F4/80，TNF-α，IL-1β and IL-6 in liver tissue. Western blot was used to detect protein expression of FXR，TLR4，MyD88 and NF-κB in liver tissue. The histopathological changes of the liver were observed by hematoxylin-eosin （HE） staining. ResultsCompared with those in the blank group，ALT，AST，ALP，γ-GT，TBA，TBil and DBil levels in serum of rats in the model group were significantly increased （P<0.01）. The levels and mRNA expression of TNF-α，IL-1β and IL-6 in liver tissue were significantly increased （P<0.01），and the mRNA and protein expressions of FXR in liver tissue were decreased （P<0.01）. The mRNA and protein expressions of TLR4，MyD88 and NF-κB and the mRNA expression of F4/80 were obviously increased （P<0.05，P<0.01）. Hepatic histopathology showed inflammatory cell infiltration and proliferative changes of bile duct epithelial cells. Compared with those in the model group，ALT，ALP，γ-GT，TBA，TBil and DBil levels in serum of rats in the ursodeoxycholic acid group were obviously decreased （P<0.05，P<0.01），and the levels and mRNA expression of TNF-α，IL-1β and IL-6 in liver tissue were obviously decreased （P<0.05，P<0.01）. The mRNA and protein expressions of TLR4，MyD88 and NF-κB and the mRNA expression of F4/80 in liver tissue were obviously decreased （P<0.05，P<0.01）. ALT，AST，ALP，γ-GT，TBA，TBil and DBil levels in the serum of rats in the Yinchenhao Tang group were obviously decreased （P<0.05，P<0.01），and the levels and mRNA expression of TNF-α，IL-1β and IL-6 in liver tissue were obviously decreased （P<0.01）. The mRNA and protein expressions of FXR in liver tissue were significantly increased，and the mRNA expressions of TLR4，MyD88，NF-κB，and F4/80， as well as the protein expressions of TLR4 and NF-κB were obviously decreased （P<0.05，P<0.01）. The inflammatory cell infiltration of liver tissue and the proliferation of bile duct epithelial cells decreased. ConclusionYinchenhao Tang has an obvious protective effect on CLI，and its mechanism may be related to regulating FXR to inhibit TLR4/MyD88/NF-κB pathway-mediated inflammatory response.

OBJECTIVE To investigate the deglycosylation metabolism of dioscin in vivo and the changes of its anti-tumor activi-ty of its deglycosylated metabolites.METHODS An LC-MS/MS analysis method for simultaneous determination of dioscin and its deglycosylation metabolites was established to study the cumulative excretion amount of dioscin and its deglycosylated metabolites in rat feces after oral administration.To mimic its deglycosylation metabolism,HPLC method was applied to investigate the time-dependent changes in the prototype components and metabolites of dioscin after incubation in artificial gastric juice.Solid-phase extraction tech-nology was employed to isolate the product of dioscin following hydrolysis by artificial gastric juice.The cytotoxic effects of this product on human colon cancer cells HCT-116 were assessed using the CCK-8 assay across different hydrolysis time periods.Concurrently,the enzymatic activities of caspase-3 and caspase-9,along with the expression levels of cytochrome C,were measured to elucidate the impact of dioscin post-hydrolysis on the cytotoxicity against HCT-116 cells.RESULTS Dioscin and its series of deglycosylated me-tabolites were detected in rat feces,revealing no significant differences in the cumulative excretion amounts of Polyphyllin Ⅴ and Pro-genin Ⅱ.Dioscin was shown to generate a range of deglycosylated metabolites in artificial gastric juice.Furthermore,dioscin and its deglycosylated metabolites inhibited the proliferation of HCT-116 cells,induced morphological changes,and increased the enzymatic activities of caspase-3 and caspase-9,as well as cytochrome C expression.However,it was observed that the antitumor activity of the deglycosylated metabolites diminished with prolonged hydrolysis time.CONCLUSION The deglycosylation metabolism of dioscin sig-nificantly attenuates its inhibitory effect on the proliferation of HCT-116 cells.Suppressing the acid-mediated or gut microbiota-medi-ated deglycosylation metabolism may be a promising strategy to preserve its antitumor activity.

Prescribing cascade is a common inappropriate drug use, which will produce and amplify the risk of polypharmacy, causing unrecognized adverse drug reactions or drug-induced diseases, but it is often ignored by clinicians and pharmacists. Prescribing cascade can not only cause harm to patients, but also lead to waste of medical resources and increases the medical burden. This paper reviews the related research progress of the prevention, identification and management of the prescribing cascade, and puts forward suggestions on the identification, termination and research methods of the prescribing cascade. Chinese medical staff should improve their understanding of prescribing cascade, strengthen relevant research, understand the current situation and characteristics of prescribing cascade in China, formulate a list of prescription cascade and study relevant management strategies.

OBJECTIVE: To retrospectively analyze the clinical characteristics, co-mutated genes in newly diagnosed acute myeloid leukemia (AML) patients with NRAS and KRAS gene mutations, and the impact of NRAS and KRAS mutations on prognosis. METHODS: The clinical data and next-generation sequencing results of 80 newly diagnosed AML patients treated at our hospital from December 2018 to December 2023 were collected. The clinical characteristics, co-mutated genes of NRAS and KRAS , and the impact of NRAS and KRAS mutations on prognosis in newly diagnosed AML patients were analyzed. RESULTS: Among 80 newly diagnosed AML patients, NRAS mutations were detected in 20 cases(25.0%), and KRAS mutations were detected in 9 cases(11.3%). NRAS mutations predominantly occurred at codons 12 and 13 of exon 2, as well as codon 61 of exon 3, while KRAS mutations were most commonly occurred at codons 12 and 13 of exon 2, all of which were missense mutations. There were no statistically significant differences observed in terms of age, sex, white blood cell count(WBC), hemoglobin(Hb), platelet count(PLT), bone marrow blasts, first induction chemotherapy regimen, CR1/CRi1 rates, chromosome karyotype, 2022 ELN risk classification and allogeneic hematopoietic stem cell transplantation(allo-HSCT) among the NRAS mutation group, KRAS mutation group and NRAS/KRAS wild-type group (P >0.05). KRAS mutations were significantly correlated with PTPN11 mutations (r =0.344), whereas no genes significantly associated with NRAS mutations were found. Survival analysis showed that compared to the NRAS/KRAS wild-type group, patients with NRAS mutation had a relatively higher 5-year overall survival (OS) rate and relapse-free survival (RFS) rate, though the differences were not statistically significant (P =0.097, P =0.249). Compared to the NRAS/KRAS wild-type group, patients with KRAS mutation had a lower 5-year OS rate and RFS rate, with no significant differences observed (P =0.275, P =0.442). There was no significant difference in the 5-year RFS rate between the KRAS mutation group and NRAS mutation group (P =0.157), but the 5-year OS rate of patients with KRAS mutation was significantly lower than that of patients with NRAS mutation (P =0.037). CONCLUSION In newly diagnosed AML patients, KRAS mutation was significantly correlated with PTPN11 mutation. Compared to patients with NRAS/KRAS wild-type, those with NRAS mutation showed a more favorable prognosis, while patients with KRAS mutation showed a poorer prognosis; however, these differences did not reach statistical significance. Notably, the prognosis of AML patients with KRAS mutation was significantly inferior compared to those with NRAS mutation.
Humans ; Leukemia, Myeloid, Acute/diagnosis* ; Mutation ; Prognosis ; Proto-Oncogene Proteins p21(ras)/genetics* ; GTP Phosphohydrolases/genetics* ; Retrospective Studies ; Membrane Proteins/genetics* ; Female ; Male ; Middle Aged ; Adult ; Aged

Lymph node metastasis (LNM) is a crucial risk factor influencing an unfavorable prognosis in specific cancers. Fundamental research illuminates our understanding of tumor behavior and identifies valuable therapeutic targets. Nevertheless, the exploration of fundamental theories and the validation of clinical therapies hinge on preclinical experiments. Preclinical models, in this context, serve as the conduit connecting fundamental theories to clinical outcomes. In vivo models established in animals offer a valuable platform for comprehensively observing interactions between tumor cells and organisms. Using various experimental animals, including mice, diverse methods, such as carcinogen-induced tumorigenesis, tumor cell line or human tumor transplantation, genetic engineering, and humanization, have been used effectively to construct numerous models for tumor LNM. Carcinogen-induced models simulate the entire process of tumorigenesis and metastasis. Transplantation models, using human tumor cell lines or patient-derived tumors, offer a research platform closely mirroring the histology and clinical behavior of human tumors. Genetically engineered models have been used to delve into the mechanisms of primary tumorigenesis within an intact microenvironment. Humanized models are used to overcome barriers between human and murine immune systems. Beyond mouse models, various other animal models have unique advantages and limitations, all contributing to exploring LNM. This review summarizes existing in vitro and animal preclinical models, identifies current bottlenecks in preclinical research, and offers an outlook on forthcoming preclinical models.
Animals ; Humans ; Mice ; Lymphatic Metastasis/pathology* ; Disease Models, Animal ; Cell Line, Tumor

OBJECTIVES: To study the therapeutic mechanism of Tuihuang Mixture against cholestasis. METHODS: Forty-eight Wistar rats were randomized equally into blank group, model group, ursodeoxycholic acid group and Tuihuang Mixture group. Except for those in the blank group, all the rats were given α‑naphthylisothiocyanate (ANIT) to establish rat models of cholestasis, followed by treatments with indicated drugs or distilled water. Serum levels of ALT, AST, ALP, γ-GT, TBA and TBIL of the rats were determined, and hepatic expressions IL-1β, IL-18, FXR, NLRP3, ASC, Caspase-1 and GSDMD were detected using q-PCR, ELISA or Western blotting. Histopathological changes of the liver tissues were observed using HE staining. RESULTS: The rat models of cholestasis had significantly increased serum levels of ALT, AST, ALP, γ-GT, TBA and TBIL with increased mRNA and protein expressions of IL-1β and IL-18, decreased protein and mRNA expressions of FXR, and increased protein expressions of NLRP3 and Caspase-1 and mRNA expressions of NLRP3, ASC, Caspase-1 and GSDMD in the liver tissue, showing also irregular arrangement of liver cells, proliferation of bile duct epithelial cells and inflammatory cells infiltration. Treatment of the rat models with Tuihuang Mixture significantly decreased serum levels of ALT, AST, ALP, γ-GT, TBA and TBIL, lowered IL-1β and IL-18 and increased FXR protein and mRNA expressions, and reduced NLRP3, ASC, Caspase-1 and GSDMD proteins and NLRP3, ASC and Caspase-1 mRNA expressions in the liver tissue. Tuihuang Mixture also significantly alleviated hepatocyte injury, bile duct epithelial cell proliferation and inflammatory cell infiltration in the liver of the rat models. CONCLUSIONS Tuihuang Mixture can effectively improve cholestasis in rats possibly by inhibiting NLRP3 inflammatosome-mediated pyroptosis via regulating FXR.
Animals ; NLR Family, Pyrin Domain-Containing 3 Protein ; Rats ; Receptors, Cytoplasmic and Nuclear/metabolism* ; Cholestasis/drug therapy* ; Rats, Wistar ; Inflammasomes/metabolism* ; 1-Naphthylisothiocyanate ; Drugs, Chinese Herbal/therapeutic use* ; Male ; Interleukin-18/metabolism* ; Caspase 1/metabolism* ; Interleukin-1beta/metabolism* ; Liver/metabolism*

Acute mitochondrial damage and the energy crisis following axonal injury highlight mitochondrial transport as an important target for axonal regeneration. Syntaphilin (Snph), known for its potent mitochondrial anchoring action, has emerged as a significant inhibitor of both mitochondrial transport and axonal regeneration. Therefore, investigating the molecular mechanisms that influence the expression levels of the snph gene can provide a viable strategy to regulate mitochondrial trafficking and enhance axonal regeneration. Here, we reveal the inhibitory effect of microRNA-146b (miR-146b) on the expression of the homologous zebrafish gene syntaphilin b (snphb). Through CRISPR/Cas9 and single-cell electroporation, we elucidated the positive regulatory effect of the miR-146b-snphb axis on Mauthner cell (M-cell) axon regeneration at the global and single-cell levels. Through escape response tests, we show that miR-146b-snphb signaling positively regulates functional recovery after M-cell axon injury. In addition, continuous dynamic imaging in vivo showed that reprogramming miR-146b significantly promotes axonal mitochondrial trafficking in the pre-injury and early stages of regeneration. Our study reveals an intrinsic axonal regeneration regulatory axis that promotes axonal regeneration by reprogramming mitochondrial transport and anchoring. This regulation involves noncoding RNA, and mitochondria-associated genes may provide a potential opportunity for the repair of central nervous system injury.
Animals ; Zebrafish ; MicroRNAs/genetics* ; Nerve Regeneration/physiology* ; Mitochondria/metabolism* ; Zebrafish Proteins/genetics* ; Axons/metabolism* ; Signal Transduction/physiology* ; Axonal Transport/physiology* ; Nerve Tissue Proteins/genetics*

Objective To establish the reference interval of urine Alzheimer-associated neuronal thread protein(AD7c-NTP)for healthy adults in Mianyang area using the indirect method.Methods The detection results of urine AD7c-NTP from 5 093 healthy in-dividuals were collected from the information management database of Medical Laboratory Department of Sichuan Science City Hospital from March 2017 to March 2022.Skewness-kurtosis and Kolmogorov-Smirnov tests were used to determine whether the data followed a normal distribution.After removing outliers using the Box Plots method,the enrolled subjects were grouped by gender and age.The Mann-Whitney U or Kruska-Wallis H tests were used to analyze the between-group differences of urine AD7c-NTP in healthy individu-als with different genders and ages.The adjacent age groups without statistically significant difference(P＞0.05)were combined,and the indirect method(non-parametric test method)was used to calculate the reference intervals for different gender and age groups.Results Skewness-kurtosis and Kolmogorov-Smirnov tests showed that the data followed a non-normal distribution.After removing 293 outliers using the Box Plots method,a total of 4 800 subjects,including 3 199 males and 1 601 females,were enrolled.The enrolled subjects were grouped by gender and age,and the non-parametric test method were used to establish the reference intervals of urine AD7c-NTP in healthy populations with different genders.The Mann-Whitney U test confirmed that urine AD7c-NTP levels existed gen-der differences(Z=14.09,P＜0.01),and the reference intervals for males and females were≤1.10 ng/mL and≤1.40 ng/mL,re-spectively.There were also statistical differences in urine AD7c-NTP levels among different age groups of the same gender.After combi-ning adjacent age groups without statistically significant difference(P＞0.05),the reference intervals of urine AD7c-NTP in healthy populations with different genders and ages were established by the non-parametric test method,which were≤1.00 ng/mL for male 20-39 years old group,≤1.10 ng/mL for male 40-79 years old group,≤1.60 ng/mL for male≥80 years old group,≤1.30 ng/mL for female 20-69 years old group,and≤1.60 ng/mL for female≥70 years old group,respectively.The established reference intervals of urine AD7c-NTP were further verified by healthy individuals,and the results met the standards.Conclusion The reference intervals of urine AD7c-NTP in healthy populations with different genders and ages in Mianyang area are established successfully using the indi-rect method,which may help to predict the risk of Alzheimer's disease in clinical practice and provide support for the diagnosis and treatment of the disease.

Aim To explore the mechanism of Cong Rong San on AD model rats based on protein kinase R-like endoplasmic reticulum kinase(PERK)-eukaryotic initiation factor 2α(eIF2α)-nuclear factor kappa B(NF-κB)signaling pathway.Methods Sixty mice were randomly divided into normal group,model group,Cong Rong San groups(4.62,9.24,18.48 g·kg-1)and donepezil group,with 10 mice in each group.All groups of rats received bilateral hippocampal injections of Aβ1-42 to establish the AD model,except the normal group.After the intragastric administration,the Morris water maze behavior test was performed for rats to test-ed the learning and memory abilities.Nissl staining was detected the quantity and Nissl bodies of nerve cells.To detect the nuclear translocation of NF-κB by immu-nofluorescence.To observe the ultrastructure of endo-plasmic reticulum by Transmission electron microsco-py.ELISA for Aβ1-42 and inflammatory cytokines quantification.Western blot was used to detect the ex-pression level of protein in the hippocampus in PERK-eIF2α-NF-κB signaling pathway.Results The morris water maze results showed that Cong Rong San im-proved the escape latency time,increased the number of platform crossings,and prolonged the time spent in the target quadrant in AD rats.(P＜0.05 or P＜0.01).Nissl staining shows the neuronal cells are ar-ranged neatly,nucleus are present and the number of Nissl bodies was numerous and the number of neurons was increased in various doses of Cong Rong San.Im-munofluorescence showed that the expression of NF-κB in the nucleus of rats was decreased(P＜0.05 or P＜0.01).The shape of endoplasmic reticulum was neat,no significantly expanded,and the structure was normal in various doses of Cong Rong San.The levels of Aβ1-42,IL-1,TNF-α and the ratio of p-PERK/PERK,p-eIF2α/eIF2α,p-NF-κB p65/NF-κB p65 in hippo-campus of Cong Rong San group was significantly de-creased in ELISA and Western blot test(P＜0.05 or P＜0.01).Conclusion Cong Rong San can alleviates the immune inflammatory response of neuronal cells in the ERS state for improve the learning and memory a-bility of AD rats,the mechanism of action may through restraint the activation of PERK-eIF2α-NF-κB signa-ling pathway.