Cognitive impairment caused by cerebral small vessel disease (CSVD) can affect the quality of life of patients and impose a huge economic burden on the families and society. Inflammation is an important pathological mechanism of cognitive impairment in CSVD and its associated cognitive impairment. This article reviews the peripheral blood inflammatory markers of cognitive impairment in patients with CSVD.

Phase Ⅰ clinical trials play a crucial role in the research and development of new drugs, serving as the initial studies to assess their safety, tolerability, effectiveness, and pharmacokinetic properties in humans. These trials involve uncertainties regarding safety and efficacy. Comprehensive management of all aspects of phase Ⅰ clinical trials for anti-tumor drugs is crucial to protect the rights and safety of participants. This article provides an in-depth analysis of the key points and precautions necessary for effective quality control throughout the process. The analysis is informed by guidelines such as the “Good Clinical Practice for Drugs” “Key Points and Judgment Principles for Drug Registration Verification” “Key Points and Judgment Principles for Supervision and Inspection of Drug Clinical Trial Institutions” and the standard operating procedures for quality control of the center. Topics discussed include informed consent, inclusion criteria, experimental drugs, biological samples, adverse events, and serious adverse events. The goal is to standardize quality control in phase Ⅰ clinical trials of anti-tumor drugs, ensure the authenticity and reliability of clinical trial data, and protect the rights and safety of participants.

Objective To quantitatively assess the correlation between the skeletal muscle index(SMI)of patients and the occur-rence of anastomotic leakage(AL)in rectal cancer patients after surgery,and to analyze the risk factors for AL in rectal cancer patients and the influencing factors of sarcopenia.Methods The clinical,pathological,and related imaging data of 362 patients who under-went radical surgery for rectal cancer were retrospectively analyzed.All patients underwent pelvic MRI and abdominal CT scans(plain/enhanced)within one month before surgery,and the third lumbar vertebra skeletal muscle area(L3-SMA)was measured from the images.All patients were divided into AL group(56 cases)and control group(306 cases)based on the presence or absence of postoperative complications.The differences in clinical characteristics and imaging parameters between the two groups were analyzed.A logistic risk prediction model was established.Results Significant differences were observed between the two groups in sarcopenia,type of surgery,surgical approach,serum albumin level,operation duration,stoma type,and extramural vascular invasion(EMVI)(P＜0.05).These factors were incorporated in a multivariate logistic regression analysis model,the area under the curve(AUC)of receiver operating characteristic(ROC)curve of the model was 0.810[95％confidence interval(CI)0.743-0.876,P＜0.001],with a sensitivity of 0.865 and specificity of 0.669.Conclusion Sar-copenia is a significant risk factor for AL after rectal cancer surgery.It enhances the predictive efficacy for postoperative AL and serves as a basis for identifying high-risk populations for AL in clinical practice.

Objective To obtain tree shrew Vasorin(VASN)recombinant protein through prokaryotic expression and purification,prepare monoclonal antibody against tree shrew VASN by immunizing mice with this protein,and preliminarily evaluate its application value.Methods Reverse transcription-polymerase chain reaction(RT-PCR)was used to amplify the full-length sequence of tree shrew VASN gene in vitro.The tree shrew VASN gene fragment was inserted into pET-30a vector to construct pET-30a-VASN recombinant plasmid.The recombinant plasmid was subjected to double digestion with BamH Ⅰ and Sal Ⅰfor identification,and its correctness was further verified by sequencing.The recombinant plasmid with correct sequencing was transformed into BL21(DE3)competent cells,and isopropyl β-D-thiogalactoside(IPTG)was used to induce expression of VASN recombinant protein.Proteins were separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis(SDS-PAGE),and the VASN recombinant protein was purified by KCI.Purified recombinant protein was used to immunize BALB/c mice for four times,and serum antibody titer was detected by enzyme-linked immunosorbent assay(ELISA).Splenocytes from mice with serum antibody titer above 1:10 000 were used for cell fusion with myeloma cells.Hypoxanthine-aminopterin-thymidine(HAT)culture medium was first used to screen hybridoma cells.ELISA was used to screen positive hybridoma cell lines that could secrete specific antibodies,and monoclonal hybridoma cell lines were obtained by limiting dilution method.VASN monoclonal antibodies were prepared in large quantities by ascites induction method,purified using rProtein G,and the affinity and in vitro reaction specificity of the monoclonal antibodies were detected by ELISA and Western blotting.Results The full-length sequence of the tree shrew VASN gene was successfully amplified and the recombinant plasmid vector of tree shrew pET-30a-VASN was constructed.The sequence obtained by sequencing of the recombinant plasmid vector was identical to the tree shrew VASN target gene sequence.Recombinant protein VASN mainly existed in the form of inclusion bodies,and the purity after purification reached 90％,meeting the requirements of subsequent immunization experiments.After four immunizations with recombinant protein VASN,mouse serum antibody titer reached 1:729 000.Monoclonal positive hybridoma cell lines were obtained through ascites induction and purification,and the constant affinity value of monoclonal antibodies measured by ELISA reached 2.59x107 L/mol.Western blotting results showed that the tree shrew VASN monoclonal antibody could bind to tree shrew VASN recombinant protein,but it showed no binding reaction with porcine retinol-binding protein 4 recombinant protein,human VASN-leucine rich repeat recombinant protein,or bovine serum albumin.Anti-tree shrew VASN monoclonal antibody could specifically recognize VASN protein in tree shrew heart,liver,spleen,lung,kidney and muscle,with clear bands and clean background.Immunohistochemical detection results showed that this monoclonal antibody could recognize VASN protein in tree shrew spleen,lung,and tree shrew immortalized fibroblasts with high VASN mRNA expression levels,and the detection results were positive.Conclusion Monoclonal antibody against tree shrew VASN is successfully prepared.This antibody can be used for immunohistochemical detection of tree shrew immortalized fibroblasts,spleen tissue,and lung tissue,providing an important tool for further research on the function of VASN in tree shrew models.

Objective To investigate the effect of Dex on neurological function in PD rats through the receptor interacting protein kinase 1(RIPK1)/RIPK3/mixed lineage kinase domain-like pro-tein(MLKL)pathway.Methods After rat PD model was constructed,the PD rats were assigned into PD group,L-,M-and H-Dex groups(intraperitoneal injection of 25,50,and 100 μg/kg Dex,respectively),and Dex+recombinant RIPK1 protein(rRIPK1)group(intraperitoneal injection of 100 μg/kg Dex+8 μg/kg rRIPK1),with 6 animals in each group.Another 6 rats served as sham-operation(Sham)group.The rats from the Sham group and PD group were intragastrically ad-ministered and injected with an equal amount of normal saline solution once a day.After 21 con-secutive days,the rotational behavior was observed of in each group rats.HE staining was per-formed to detect the pathological changes in dopaminergic neurons in the substantia nigra.TUNEL staining was conducted to measure the apoptosis of dopaminergic neurons.ELISA was used to determine the levels of neurotransmitters[DA,HVA,3,4-dihydroxyphenylacetic acid(DOPAC),5-hydroxytrptamine(5-HT)]and inflammatory factors(TNF-α,IL-1β)in the sub-stantia nigra.Western blotting was applied to detect the expression of RIPK1/RIPK3/MLKL pathway related proteins in the substantia nigra tissues.Results Intact and well-arranged mor-phology and structure were observed in the neurons of the Sham group.The PD group presented prominently less neurons,in scattered arrangement,with obviously reduced volume,irregular nu-clear deformation,indicating notably neuronal damage.The severity of neuronal damage was at-tenuated sequentially in the L-Dex,M-Dex,and H-Dex groups,but the damage in the Dex+rRIPK1 group was further worsened.The PD group had significantly larger number of rotations,longer escape latency,higher apoptotic rate,increased TNF-α and IL-1β contents,and elevated lev-els of p-RIPK1/RIPK1(1.07±0.18 vs 0.36±0.11),p-RIPK3/RIPK3(1.32±0.21 vs 0.47±0.14),and p-MLKL/MLKL(0.79±0.11 vs 0.18±0.05),but lower DA,DOPAC,5-HT,and HVA con-tents than the Sham group(P＜0.05).Dex treatment of low,medium and high doses reserved all above changes induced by PD modeling(P＜0.05).The Dex+rRIPK1 group obtained larger num-ber of rotations,longer escape latency,higher apoptotic rate,increased TNF-α and IL-1β contents,and elevated levels of p-RIPK1/RIPK1(0.95±0.17 vs 0.41±0.12),p-RIPK3/RIPK3(1.14±0.20 vs 0.51±0.15),and p-MLKL/MLKL(0.72±0.09 vs 0.24±0.06),but decreased DA,DOPAC,5-HT,and HVA contents when compared with the H-Dex group(P＜0.05).Conclusion Dex protects the neurological function of PD rats by inhibiting the RIPK1/RIPK3/MLKL pathway.

Objective:To improve the diagnosis and treatment of autoimmune hepatitis (AIH) in children by analyzing their clinical characteristics and outcomes.Methods:Case summary.The general data, clinical manifestations, laboratory parameters, liver pathology, treatment and outcomes of 6 AIH children diagnosed at Xi′an Children′s Hospital from January 2019 to March 2024 were retrospectively analyzed.Results:All the 6 patients were diagnosed with AIH-1, and 5 of them were female, accounting for 83.3%.The median age of onset was 6.2 years old.Three cases (50.0%) had insidious onset.Elevated hepatic aminotransferase levels were detected incidentally in 2 cases (33.3%).One case (16.7%) presented symptoms resembling those of viral hepatitis.Two cases (33.3%) were accompanied by extrahepatic autoimmune diseases.All the children tested positive for anti-nuclear antibodies with a titer ≥1∶80 or 1∶100.Four cases (66.7%), 3 cases (50.0%), and 1 case (16.7%) tested positive for anti-smooth muscle antibodies, anti-perinuclear neutrophil cytoplasm antibodies and anti-soluble liver antigen, respectively.Liver pathology showed infiltration of lymphocytes-plasma cells and moderate or severe interfacial inflammation in all the 6 cases.Two cases (33.3%) had rosettes.All the patients were given oral corticosteroids combined with Azathioprine or Mycophenolate mofetil.The patients were followed up for 24.8 (12.0-52.0) months, during which 3 cases (50.0%) had partial remission and 3 cases (50.0%) achieved complete remission.Four cases (66.7%) showed transaminase fluctuations during corticosteroid reduction or cessation.Conclusions:Autoantibodies and liver pathology are important evidence for diagnosing AIH in children.Immunosuppressants have a high remission rate in the treatment of this disease, but transaminase fluctuations are prone to occur during corticosteroid reduction or cessation.

Objective:To improve the diagnosis and treatment of autoimmune hepatitis (AIH) in children by analyzing their clinical characteristics and outcomes.Methods:Case summary.The general data, clinical manifestations, laboratory parameters, liver pathology, treatment and outcomes of 6 AIH children diagnosed at Xi′an Children′s Hospital from January 2019 to March 2024 were retrospectively analyzed.Results:All the 6 patients were diagnosed with AIH-1, and 5 of them were female, accounting for 83.3%.The median age of onset was 6.2 years old.Three cases (50.0%) had insidious onset.Elevated hepatic aminotransferase levels were detected incidentally in 2 cases (33.3%).One case (16.7%) presented symptoms resembling those of viral hepatitis.Two cases (33.3%) were accompanied by extrahepatic autoimmune diseases.All the children tested positive for anti-nuclear antibodies with a titer ≥1∶80 or 1∶100.Four cases (66.7%), 3 cases (50.0%), and 1 case (16.7%) tested positive for anti-smooth muscle antibodies, anti-perinuclear neutrophil cytoplasm antibodies and anti-soluble liver antigen, respectively.Liver pathology showed infiltration of lymphocytes-plasma cells and moderate or severe interfacial inflammation in all the 6 cases.Two cases (33.3%) had rosettes.All the patients were given oral corticosteroids combined with Azathioprine or Mycophenolate mofetil.The patients were followed up for 24.8 (12.0-52.0) months, during which 3 cases (50.0%) had partial remission and 3 cases (50.0%) achieved complete remission.Four cases (66.7%) showed transaminase fluctuations during corticosteroid reduction or cessation.Conclusions:Autoantibodies and liver pathology are important evidence for diagnosing AIH in children.Immunosuppressants have a high remission rate in the treatment of this disease, but transaminase fluctuations are prone to occur during corticosteroid reduction or cessation.

Objective To obtain tree shrew Vasorin(VASN)recombinant protein through prokaryotic expression and purification,prepare monoclonal antibody against tree shrew VASN by immunizing mice with this protein,and preliminarily evaluate its application value.Methods Reverse transcription-polymerase chain reaction(RT-PCR)was used to amplify the full-length sequence of tree shrew VASN gene in vitro.The tree shrew VASN gene fragment was inserted into pET-30a vector to construct pET-30a-VASN recombinant plasmid.The recombinant plasmid was subjected to double digestion with BamH Ⅰ and Sal Ⅰfor identification,and its correctness was further verified by sequencing.The recombinant plasmid with correct sequencing was transformed into BL21(DE3)competent cells,and isopropyl β-D-thiogalactoside(IPTG)was used to induce expression of VASN recombinant protein.Proteins were separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis(SDS-PAGE),and the VASN recombinant protein was purified by KCI.Purified recombinant protein was used to immunize BALB/c mice for four times,and serum antibody titer was detected by enzyme-linked immunosorbent assay(ELISA).Splenocytes from mice with serum antibody titer above 1:10 000 were used for cell fusion with myeloma cells.Hypoxanthine-aminopterin-thymidine(HAT)culture medium was first used to screen hybridoma cells.ELISA was used to screen positive hybridoma cell lines that could secrete specific antibodies,and monoclonal hybridoma cell lines were obtained by limiting dilution method.VASN monoclonal antibodies were prepared in large quantities by ascites induction method,purified using rProtein G,and the affinity and in vitro reaction specificity of the monoclonal antibodies were detected by ELISA and Western blotting.Results The full-length sequence of the tree shrew VASN gene was successfully amplified and the recombinant plasmid vector of tree shrew pET-30a-VASN was constructed.The sequence obtained by sequencing of the recombinant plasmid vector was identical to the tree shrew VASN target gene sequence.Recombinant protein VASN mainly existed in the form of inclusion bodies,and the purity after purification reached 90％,meeting the requirements of subsequent immunization experiments.After four immunizations with recombinant protein VASN,mouse serum antibody titer reached 1:729 000.Monoclonal positive hybridoma cell lines were obtained through ascites induction and purification,and the constant affinity value of monoclonal antibodies measured by ELISA reached 2.59x107 L/mol.Western blotting results showed that the tree shrew VASN monoclonal antibody could bind to tree shrew VASN recombinant protein,but it showed no binding reaction with porcine retinol-binding protein 4 recombinant protein,human VASN-leucine rich repeat recombinant protein,or bovine serum albumin.Anti-tree shrew VASN monoclonal antibody could specifically recognize VASN protein in tree shrew heart,liver,spleen,lung,kidney and muscle,with clear bands and clean background.Immunohistochemical detection results showed that this monoclonal antibody could recognize VASN protein in tree shrew spleen,lung,and tree shrew immortalized fibroblasts with high VASN mRNA expression levels,and the detection results were positive.Conclusion Monoclonal antibody against tree shrew VASN is successfully prepared.This antibody can be used for immunohistochemical detection of tree shrew immortalized fibroblasts,spleen tissue,and lung tissue,providing an important tool for further research on the function of VASN in tree shrew models.

Objective To quantitatively assess the correlation between the skeletal muscle index(SMI)of patients and the occur-rence of anastomotic leakage(AL)in rectal cancer patients after surgery,and to analyze the risk factors for AL in rectal cancer patients and the influencing factors of sarcopenia.Methods The clinical,pathological,and related imaging data of 362 patients who under-went radical surgery for rectal cancer were retrospectively analyzed.All patients underwent pelvic MRI and abdominal CT scans(plain/enhanced)within one month before surgery,and the third lumbar vertebra skeletal muscle area(L3-SMA)was measured from the images.All patients were divided into AL group(56 cases)and control group(306 cases)based on the presence or absence of postoperative complications.The differences in clinical characteristics and imaging parameters between the two groups were analyzed.A logistic risk prediction model was established.Results Significant differences were observed between the two groups in sarcopenia,type of surgery,surgical approach,serum albumin level,operation duration,stoma type,and extramural vascular invasion(EMVI)(P＜0.05).These factors were incorporated in a multivariate logistic regression analysis model,the area under the curve(AUC)of receiver operating characteristic(ROC)curve of the model was 0.810[95％confidence interval(CI)0.743-0.876,P＜0.001],with a sensitivity of 0.865 and specificity of 0.669.Conclusion Sar-copenia is a significant risk factor for AL after rectal cancer surgery.It enhances the predictive efficacy for postoperative AL and serves as a basis for identifying high-risk populations for AL in clinical practice.

Objective To investigate the effect of Dex on neurological function in PD rats through the receptor interacting protein kinase 1(RIPK1)/RIPK3/mixed lineage kinase domain-like pro-tein(MLKL)pathway.Methods After rat PD model was constructed,the PD rats were assigned into PD group,L-,M-and H-Dex groups(intraperitoneal injection of 25,50,and 100 μg/kg Dex,respectively),and Dex+recombinant RIPK1 protein(rRIPK1)group(intraperitoneal injection of 100 μg/kg Dex+8 μg/kg rRIPK1),with 6 animals in each group.Another 6 rats served as sham-operation(Sham)group.The rats from the Sham group and PD group were intragastrically ad-ministered and injected with an equal amount of normal saline solution once a day.After 21 con-secutive days,the rotational behavior was observed of in each group rats.HE staining was per-formed to detect the pathological changes in dopaminergic neurons in the substantia nigra.TUNEL staining was conducted to measure the apoptosis of dopaminergic neurons.ELISA was used to determine the levels of neurotransmitters[DA,HVA,3,4-dihydroxyphenylacetic acid(DOPAC),5-hydroxytrptamine(5-HT)]and inflammatory factors(TNF-α,IL-1β)in the sub-stantia nigra.Western blotting was applied to detect the expression of RIPK1/RIPK3/MLKL pathway related proteins in the substantia nigra tissues.Results Intact and well-arranged mor-phology and structure were observed in the neurons of the Sham group.The PD group presented prominently less neurons,in scattered arrangement,with obviously reduced volume,irregular nu-clear deformation,indicating notably neuronal damage.The severity of neuronal damage was at-tenuated sequentially in the L-Dex,M-Dex,and H-Dex groups,but the damage in the Dex+rRIPK1 group was further worsened.The PD group had significantly larger number of rotations,longer escape latency,higher apoptotic rate,increased TNF-α and IL-1β contents,and elevated lev-els of p-RIPK1/RIPK1(1.07±0.18 vs 0.36±0.11),p-RIPK3/RIPK3(1.32±0.21 vs 0.47±0.14),and p-MLKL/MLKL(0.79±0.11 vs 0.18±0.05),but lower DA,DOPAC,5-HT,and HVA con-tents than the Sham group(P＜0.05).Dex treatment of low,medium and high doses reserved all above changes induced by PD modeling(P＜0.05).The Dex+rRIPK1 group obtained larger num-ber of rotations,longer escape latency,higher apoptotic rate,increased TNF-α and IL-1β contents,and elevated levels of p-RIPK1/RIPK1(0.95±0.17 vs 0.41±0.12),p-RIPK3/RIPK3(1.14±0.20 vs 0.51±0.15),and p-MLKL/MLKL(0.72±0.09 vs 0.24±0.06),but decreased DA,DOPAC,5-HT,and HVA contents when compared with the H-Dex group(P＜0.05).Conclusion Dex protects the neurological function of PD rats by inhibiting the RIPK1/RIPK3/MLKL pathway.