Objective To investigate the characteristics of liver injury in the Lieber-DeCarli alcoholic liver disease(ALD)mouse model and to analyze its transcriptomic profile.Methods Eighteen male C57BL/6J mice were randomly divided into an alcohol-fed group(n = 10)and a control group(n = 8).The alcohol-fed group received a Lieber-DeCarli ethanol diet,starting with an adaptive one-week phase using incremental concentrations of ethanol(10～57.3 mL/L),followed by 2 weeks of a 57.3 mL/L concentration of 95％ethanol,for a total of 3 weeks.The control group was provided with an isocaloric control diet for 3 weeks.At the end of the study,mice were sacrificed,and serum and liver tissue samples were collected.Serum liver function markers(ALT,AST),hepatic lipids(TC,TG),reduced glutathione(GSH),total superoxide dismutase(T-SOD),and malondialdehyde(MDA)were measured using biochemical assays.The levels of inflammatory cytokines(IL-6,IL-10,TNF-α,TGF-β1)in liver tissue were assessed by ELISA.Histopathological changes in liver tissue were examined using hematoxylin-eosin(HE)and Oil Red O staining.Immunohistochemical staining using the F4/80 antibody was employed to assess changes in macrophage expression.RNA-seq analysis was conducted to identify differentially expressed genes between the two groups of liver tissues,followed by GO and KEGG pathway enrichment analysis.qRT-PCR was used to validate the expression of these differentially expressed genes.Results Compared with the control group,the alcohol-fed mice exhibited a significant decrease in body weight(P<0.01).Serum ALT and AST levels were significantly elevated(P<0.01),while liver tissue levels of TC,TG,and MDA were significantly increased(P<0.05).Conversely,GSH and T-SOD levels were significantly reduced(P<0.05).The levels of inflammatory factors IL-6,TNF-α,and TGF-β1 were increased,which was consistent with the qRT-PCR validation results(P<0.05).Histological examination revealed disrupted hepatic lobular structure,with macrovesicular steatosis,microvesicular steatosis,and ballooning degeneration.Additionally,fat droplets in liver tissue were significantly increased,and macrophage expression was upregulated.Differential gene expression analysis,using a threshold of|log2 FC|>1 and q<0.05,identified 2063 differentially expressed genes,of which 1236 were upregulated and 827 downregulated.Enriched pathways included xenobiotic metabolism via cytochrome P450,cytokine-cytokine receptor interaction,chemokine signaling,steroid hormone biosynthesis,glutathione metabolism,and retinol metabolism.(P<0.05).qRT-PCR validation confirmed the significant upregulation(e.g.,Mmp12,Gstm3,Cyp2a22)and downregulation(e.g.,Serpina1e,Acmsd,Mup3d)of 10 genes from each category,consistent with the transcriptome sequencing results.Conclusions The primary pathological mechanisms underlying alcoholic liver injury involve pathways related to xenobiotic metabolism and act via cytochrome P450,cytokine-cytokine receptor interaction,chemokine signaling,glutathione metabolism,and retinol metabolism.

Objective To investigate the characteristics of liver injury in the Lieber-DeCarli alcoholic liver disease(ALD)mouse model and to analyze its transcriptomic profile.Methods Eighteen male C57BL/6J mice were randomly divided into an alcohol-fed group(n = 10)and a control group(n = 8).The alcohol-fed group received a Lieber-DeCarli ethanol diet,starting with an adaptive one-week phase using incremental concentrations of ethanol(10～57.3 mL/L),followed by 2 weeks of a 57.3 mL/L concentration of 95％ethanol,for a total of 3 weeks.The control group was provided with an isocaloric control diet for 3 weeks.At the end of the study,mice were sacrificed,and serum and liver tissue samples were collected.Serum liver function markers(ALT,AST),hepatic lipids(TC,TG),reduced glutathione(GSH),total superoxide dismutase(T-SOD),and malondialdehyde(MDA)were measured using biochemical assays.The levels of inflammatory cytokines(IL-6,IL-10,TNF-α,TGF-β1)in liver tissue were assessed by ELISA.Histopathological changes in liver tissue were examined using hematoxylin-eosin(HE)and Oil Red O staining.Immunohistochemical staining using the F4/80 antibody was employed to assess changes in macrophage expression.RNA-seq analysis was conducted to identify differentially expressed genes between the two groups of liver tissues,followed by GO and KEGG pathway enrichment analysis.qRT-PCR was used to validate the expression of these differentially expressed genes.Results Compared with the control group,the alcohol-fed mice exhibited a significant decrease in body weight(P<0.01).Serum ALT and AST levels were significantly elevated(P<0.01),while liver tissue levels of TC,TG,and MDA were significantly increased(P<0.05).Conversely,GSH and T-SOD levels were significantly reduced(P<0.05).The levels of inflammatory factors IL-6,TNF-α,and TGF-β1 were increased,which was consistent with the qRT-PCR validation results(P<0.05).Histological examination revealed disrupted hepatic lobular structure,with macrovesicular steatosis,microvesicular steatosis,and ballooning degeneration.Additionally,fat droplets in liver tissue were significantly increased,and macrophage expression was upregulated.Differential gene expression analysis,using a threshold of|log2 FC|>1 and q<0.05,identified 2063 differentially expressed genes,of which 1236 were upregulated and 827 downregulated.Enriched pathways included xenobiotic metabolism via cytochrome P450,cytokine-cytokine receptor interaction,chemokine signaling,steroid hormone biosynthesis,glutathione metabolism,and retinol metabolism.(P<0.05).qRT-PCR validation confirmed the significant upregulation(e.g.,Mmp12,Gstm3,Cyp2a22)and downregulation(e.g.,Serpina1e,Acmsd,Mup3d)of 10 genes from each category,consistent with the transcriptome sequencing results.Conclusions The primary pathological mechanisms underlying alcoholic liver injury involve pathways related to xenobiotic metabolism and act via cytochrome P450,cytokine-cytokine receptor interaction,chemokine signaling,glutathione metabolism,and retinol metabolism.

Warthin-like papillary renal cell carcinoma (WPRCC), which is rare in clinical practice, is a new subtype of papillary renal cell carcinoma in the 5th editon WHO Classification of Tumors of the Urinary and Male Reproductive System. In this paper, we report one case of WPRCC. The patient was admitted to the hospital due to a mass in the right kidney which was detected by magnetic resonance enhancement scan in physical examination. Subsequently, the laparoscopic partial nephrectomy was performed and the diagnosis of Warthin-like papillary renal cell carcinoma was confirmed. The patient was treated with interferon, and no recurrence and metastasis were found in the imaging examination after 7 months of follow-up.

Warthin-like papillary renal cell carcinoma (WPRCC), which is rare in clinical practice, is a new subtype of papillary renal cell carcinoma in the 5th editon WHO Classification of Tumors of the Urinary and Male Reproductive System. In this paper, we report one case of WPRCC. The patient was admitted to the hospital due to a mass in the right kidney which was detected by magnetic resonance enhancement scan in physical examination. Subsequently, the laparoscopic partial nephrectomy was performed and the diagnosis of Warthin-like papillary renal cell carcinoma was confirmed. The patient was treated with interferon, and no recurrence and metastasis were found in the imaging examination after 7 months of follow-up.

Objective Transcriptome sequencing technology(RNA-seq)was used to analyze the mechanism of compound Dancao granules as an intervention for high-fat feed combined with carbon tetrachloride(CCl4)-induced non-alcoholic steatohepatitis.Methods 45 male C57BL/6J mice were split into two groups at random:normal control group,model control group,obeccholic acid group 10 mg/(kg·d),and compound Dancao granules low-and high-dose groups 3.74 g/(kg·d)and 7.48 g/(kg·d),with 9 mice in each group.Normal diet was made available to the control group,and the mice in the model group were given a high-fat diet combined with the subcutaneous injection of CC14,with 100％CC14 solution(4 mL/kg)in the first application,and 40％CC14-olive oil solution(2 mL/kg)in the second application,twice a week for a total of 6 weeks.Each drug group was administered the respective drug from week 3 for a total of 4 weeks.12 h after the last administration,the serum and liver tissues of mice in each group were collected,and a biochemical kit was used to detect serum liver function.Hematoxylin-eosin(HE),sirius scarlet,and oil red O staining were used to examine histopathological changes to the liver.The levels of IL-6,IL-10,TNF-α and TGF-β in mice liver were detected via ELISA,and the expression of α-SMA was observed by immunohistochemistry.Differential gene expression was analyzed by RNA-seq and functional enrichment analysis.To verify the differential expression of mRNA,quantitative reverse transcription PCR(qRT-PCR)was used.TDT-mediated dUTP nick-end labeling(TUNEL)staining was employed to identify apoptosis.Results The model control groups had significantly higher levels of serum alanine aminotransferase(ALT),aspartate aminotransferase(AST),total cholesterol(TC),and triglycerides(TG)than normal control group(P＜0.01).Additionally,there was obvious inflammatory cell infiltration in the liver tissue,collagen deposition in the sink and interlobule areas,and a significant increase in lipid droplet area(P＜0.01).The levels of IL-6 and TNF-α in liver tissue were significantly increased(P＜0.01),the levels of IL-10 and TGF-β were decreased(P＜0.01),and the expression of α-SMA was significantly increased(P＜0.01).The levels of TC,TG,ALT,and AST were significantly lower in groups that received compound Dancao granules and obeccholic acid than the model control group(P＜0.01),and inflammatory cell infiltration,collagen deposition,and fat accumulation in the sink and interlobule areas were improved(P＜0.01).The levels of IL-6 and TNF-α in liver tissue were significantly decreased(P＜0.01),the levels of IL-10 and TGF-β were increased(P＜0.05,P＜0.01),and the expression of α-SMA was significantly decreased(P＜0.01).RNA-seq sequencing result showed that 2819 genes in the normal control group were differentially expressed compared with the model control group,with 543 up-regulated and 2276 down-regulated genes.In a comparison of the model control group and compound Dancao granules group,240 genes were differentially expressed,including 206 up-regulated genes and 34 down-regulated genes.There were 221 genes with overlapping expression in the 2 groups and functional enrichment highlighted cell cycle(Cdt1,Plk1,Bub1b,Ttk,Knl1,Esco2,Cdc6,Ndc80,Cdc25b,Sgo1,Ccnb2,Espl1,Ccne1,Mcm4,Mcm5,Fbxo5,Bub1,Mcm2),apoptosis(Caspase3,Bax,P53,Apaf1,Bak,Caspase8),the P53 signaling pathway(P53,Ccnb2,Apaf1,Bak,Bax,Gtse1,Caspase3,Ccne1),arachidonic acid metabolism(Hpgds,Cyp2c54,Cyp2b10,Tbxas1,Cyp2c50),galactose metabolism(Hk3,Gla,Hk2,Akr1b7)and other signaling pathway genes.RNA-seq sequencing analysis showed that compound Danicao granules mainly regulated the apoptosis signaling pathway,and qRT-PCR confirmed that the mRNA expression of Caspase3,Bax,P53,Apaf1,Bak and Caspase8 in the liver tissue of the model control group was increased compared with that of the normal control group(P＜0.01).Compared with the model control group,the compound Dancao granules group showed decreased mRNA expression of Caspase3,Bax,P53,Apaf1,Bak and Caspase8 in liver tissue(P＜0.01).TUNEL staining showed that the number of cells showing nuclear shrinkage and apoptotic bodies decreased in the compound Dancao granule administration group.Conclusions Compound Dancao granules had a significant protective effect against non-alcoholic steatohepatitis induced by high-fat feed combined with CCl4,and its mechanism might be connected to the control of genes linked to apoptosis.

Objective Transcriptome sequencing technology(RNA-seq)was used to analyze the mechanism of compound Dancao granules as an intervention for high-fat feed combined with carbon tetrachloride(CCl4)-induced non-alcoholic steatohepatitis.Methods 45 male C57BL/6J mice were split into two groups at random:normal control group,model control group,obeccholic acid group 10 mg/(kg·d),and compound Dancao granules low-and high-dose groups 3.74 g/(kg·d)and 7.48 g/(kg·d),with 9 mice in each group.Normal diet was made available to the control group,and the mice in the model group were given a high-fat diet combined with the subcutaneous injection of CC14,with 100％CC14 solution(4 mL/kg)in the first application,and 40％CC14-olive oil solution(2 mL/kg)in the second application,twice a week for a total of 6 weeks.Each drug group was administered the respective drug from week 3 for a total of 4 weeks.12 h after the last administration,the serum and liver tissues of mice in each group were collected,and a biochemical kit was used to detect serum liver function.Hematoxylin-eosin(HE),sirius scarlet,and oil red O staining were used to examine histopathological changes to the liver.The levels of IL-6,IL-10,TNF-α and TGF-β in mice liver were detected via ELISA,and the expression of α-SMA was observed by immunohistochemistry.Differential gene expression was analyzed by RNA-seq and functional enrichment analysis.To verify the differential expression of mRNA,quantitative reverse transcription PCR(qRT-PCR)was used.TDT-mediated dUTP nick-end labeling(TUNEL)staining was employed to identify apoptosis.Results The model control groups had significantly higher levels of serum alanine aminotransferase(ALT),aspartate aminotransferase(AST),total cholesterol(TC),and triglycerides(TG)than normal control group(P＜0.01).Additionally,there was obvious inflammatory cell infiltration in the liver tissue,collagen deposition in the sink and interlobule areas,and a significant increase in lipid droplet area(P＜0.01).The levels of IL-6 and TNF-α in liver tissue were significantly increased(P＜0.01),the levels of IL-10 and TGF-β were decreased(P＜0.01),and the expression of α-SMA was significantly increased(P＜0.01).The levels of TC,TG,ALT,and AST were significantly lower in groups that received compound Dancao granules and obeccholic acid than the model control group(P＜0.01),and inflammatory cell infiltration,collagen deposition,and fat accumulation in the sink and interlobule areas were improved(P＜0.01).The levels of IL-6 and TNF-α in liver tissue were significantly decreased(P＜0.01),the levels of IL-10 and TGF-β were increased(P＜0.05,P＜0.01),and the expression of α-SMA was significantly decreased(P＜0.01).RNA-seq sequencing result showed that 2819 genes in the normal control group were differentially expressed compared with the model control group,with 543 up-regulated and 2276 down-regulated genes.In a comparison of the model control group and compound Dancao granules group,240 genes were differentially expressed,including 206 up-regulated genes and 34 down-regulated genes.There were 221 genes with overlapping expression in the 2 groups and functional enrichment highlighted cell cycle(Cdt1,Plk1,Bub1b,Ttk,Knl1,Esco2,Cdc6,Ndc80,Cdc25b,Sgo1,Ccnb2,Espl1,Ccne1,Mcm4,Mcm5,Fbxo5,Bub1,Mcm2),apoptosis(Caspase3,Bax,P53,Apaf1,Bak,Caspase8),the P53 signaling pathway(P53,Ccnb2,Apaf1,Bak,Bax,Gtse1,Caspase3,Ccne1),arachidonic acid metabolism(Hpgds,Cyp2c54,Cyp2b10,Tbxas1,Cyp2c50),galactose metabolism(Hk3,Gla,Hk2,Akr1b7)and other signaling pathway genes.RNA-seq sequencing analysis showed that compound Danicao granules mainly regulated the apoptosis signaling pathway,and qRT-PCR confirmed that the mRNA expression of Caspase3,Bax,P53,Apaf1,Bak and Caspase8 in the liver tissue of the model control group was increased compared with that of the normal control group(P＜0.01).Compared with the model control group,the compound Dancao granules group showed decreased mRNA expression of Caspase3,Bax,P53,Apaf1,Bak and Caspase8 in liver tissue(P＜0.01).TUNEL staining showed that the number of cells showing nuclear shrinkage and apoptotic bodies decreased in the compound Dancao granule administration group.Conclusions Compound Dancao granules had a significant protective effect against non-alcoholic steatohepatitis induced by high-fat feed combined with CCl4,and its mechanism might be connected to the control of genes linked to apoptosis.

ObjectiveTo establish a polymerase chain reaction（PCR） method to accurately discriminate the crude materials of Murrayae Folium et Cacumen， Murraya exotica and M. paniculata. MethodBased on the difference in chloroplast genome sequences of M. exotica and M. paniculata， species-specific identification primers P03 and P04 of M. exotica and M. paniculata were designed according to single nucleotide polymorphism （SNP） on the chloroplast genome. A multiplex allele-specific PCR identification method was established for the identification of M. exotica and M. paniculata following the optimization of annealing temperature， number of cycles， and primer concentration ratio. The established PCR method for identification was explored and verified in terms of tolerance and feasibility by investigating the type of Taq polymerases and PCR system model. ResultIn this multiplex allele-specific PCR identification method， about 330 and 230 bp of specific fragments were amplified from DNA templates of M. exotica and M. paniculata， respectively， under the following conditions：cycle number of 31， annealing temperature of 60 ℃， and primer concentration ratio of P03 and P04 of 1∶2. Consistent results were obtained for samples from different sources. ConclusionThe multiplex allele-specific PCR identification method established in this study can accurately identify the origin of Murrayae Folium et Cacumen， which can be used for the simultaneous identification of M. exotica and M. paniculata by the length of fragments in a single identification assay.

Objective To investigate the instant clinical efficacy of intra-arterial infusion of fasudil combined with routine anti-vasospasm for symptomatic cerebral vasospasm (SCVS). Methods The clinical data of 21 patients with subarachnoid hemorrhage (SAH) due to ruptured aneurysm, who were admitted to authors' hospital during the period from May 2010 and February 2014, were retrospectively analyzed. The lesions included Fisher gradeⅡ(n=2), gradeⅢ (n=16) and gradeⅣ (n=3). Endovascular embolization of the aneurysm was carried out within 48 hours after the confirmation of the diagnosis with total cerebral DSA;no bleeding occurred during the operation and routine anti-vasospasm therapy was given. Within 4-9 days after the onset of the disease, all 21 patients presented SCVS. Half dose systemic heparinization, superselective intra-arterial infusion of fasudil (30 mg fasudil+250 ml saline, lasting for 30 min) were adopted. Reexamination of angiography performed at 15 min after fasudil infusion was employed, and the results were evaluated with NIHSS score by comparing the preoperative findings. Results Imaging examination performed after the treatment showed that significant improvement was obtained in 15 patients and no obvious changes in 6 patients. Clinical symptoms were remarkably improved in 11 patients, partially improved in 4 patients and remained unchanged in 6 patients. The mean NIHSS score was improved from preoperative 28.6 to postoperative 21.2. Conclusion For the treatment of symptomatic cerebral vasospasm, superselective intra-arterial infusion of fasudil is effective and safe, and it has good clinical application value.