ObjectiveTo investigate the effect of transplantation of bone marrow mesenchymal stem cells （BMSCs） co-cultured with bone marrow-derived M2 macrophages （M2-BMDMs）， named as BMSC^M2， on a rat model of liver cirrhosis induced by carbon tetrachloride （CCl₄）/2-acetaminofluorene （2-AAF）. MethodsRat BMDMs were isolated and polarized into M2 phenotype， and rat BMSCs were isolated and co-cultured with M2-BMDMs at the third generation to obtain BMSC^M2. The rats were given subcutaneous injection of CCl₄ for 6 weeks to establish a model of liver cirrhosis， and then they were randomly divided into model group （M group）， BMSC group， and BMSC^M2 group， with 6 rats in each group. A normal group （N group） with 6 rats was also established. Since week 7， the model rats were given 2-AAF by gavage in addition to the subcutaneous injection of CCl₄. Samples were collected at the end of week 10 to observe liver function， liver histopathology， and hydroxyproline （Hyp） content in liver tissue， as well as changes in the markers for hepatic stellate cells， hepatic progenitor cells， cholangiocytes， and hepatocytes. A one-way analysis of variance was used for comparison of continuous data between multiple groups， and the least significant difference t-test was used for further comparison between two groups. ResultsCompared with the N group， the M group had significant increases in the activities of serum alanine aminotransferase （ALT） and aspartate aminotransferase （AST） （P<0.01）； compared with the M group， the BMSC and BMSC^M2 groups had significant reductions in ALT and AST （P<0.01）， and the BMSC^M2 group had significantly better activities than the BMSC group （P<0.05）. Compared with the N group， the M group had significant increases in Hyp content and the mRNA and protein expression levels of alpha-smooth muscle actin （α-SMA） in the liver （P<0.01）； compared with the M group， the BMSC and BMSC^M2 groups had significant reductions in Hyp content and the expression of α-SMA （P<0.05）， and the BMSC^M2 group had a significantly lower level of α-SMA than the BMSC group （P<0.01）. Compared with the N group， the M group had significant increases in the mRNA expression levels of the hepatic progenitor cell markers EpCam and Sox9 and the cholangiocyte markers CK7 and CK19 （P<0.01） and significant reductions in the expression levels of the hepatocyte markers HNF-4α and Alb （P<0.01）； compared with the M group， the BMSC and BMSC^M2 groups had significant reductions in the mRNA expression levels of EpCam， Sox9， CK7， and CK19 （P<0.05） and significant increases in the mRNA expression levels of HNF-4α and Alb （P<0.05）， and compared with the BMSC group， the BMSC^M2 group had significant reductions in the mRNA expression levels of EpCam and CK19 （P<0.05） and significant increase in the expression level of HNF-4α （P<0.05）. ConclusionM2-BMDMs can enhance the therapeutic effect of BMSCs on CCl₄/2-AAF-induced liver cirrhosis in rats， which provides new ideas for further improving the therapeutic effect of BMSCs on liver cirrhosis.

Objective To investigate the clinicopathological features and key points of diagnosis and treatment of malignant perivascular epithelioid cell tumor(PEComa)to increase awareness of the disease.Methods The clinicopathological data of 2 patients with malignant PEComa treated in our hospital were retrospectively analyzed,and relevant literatures were reviewed.Results Both patients were male,aged 53 and 16 years,respectively.The sites of occurrence were in the retroperitoneum and pelvis,respectively.Both tumors were resected surgically,and the diagnosis was confirmed with postoperative pathology.Under the microscope,the tumor tissue of one patient was mainly composed of smooth muscle-like cells,and that of the other patient was composed of epithelioid cells,both showing pathological mitotic images and expressing HMB45,Melan-A,SMA and CD34,no tumor recurrence or metastasis was observed during the follow-up.The literatures collected involved 15 patients with retroperitoneal or pelvic PEComa,including 3 males and 12 females,of which 9 were malignant.The clinical manifestations were abdominal pain,bloating,or lower back pain.Some cases were detected during physical examinations.Conclusion Malignant PEComa is difficult to be diagnosed before surgery and easy to be misdiagnosed.The confirmed diagnosis depends on the postoperative pathological results.The preferred treatment is complete resection of tumor.Long-term follow-up is needed.

Objective To explore the risk factors that affect the outcome of testicular torsion in patients with testicular resection,to provide ideas for clinical management.Methods The clinical data of 117 patients with testicular torsion treated in our hospital during Jan.1,2012 and Mar.1,2023 were retrospectively analyzed,43 of whom underwent testicular detorsion and orchidopexy,and 74 underwent orchiectomy.The correlation between preoperative inflammatory and coagulation-related indicators,testicular torsion angle,TWIST score and orchiectomy was analyzed with univariate analysis.The independent risk factors for testicular torsion patients with testicular resection as the outcome were analyzed with binary logistic regression.The predictive value of each factor for orchiectomy was assessed with receiver operating characteristic(ROC)curve.Results Binary logistic regression analysis indicated that long duration of onset(OR=1.841,P=0.036),large torsion angle(OR=1.005,P=0.016),high TWIST score(OR=2.225,P=0.003)and elevated FIB level(OR=2.489,P=0.049)were independent risk factors for testicular torsion patients with testicular resection as the outcome.ROC curve analysis showed that the best cut-off value of the above 4 factors to predict orchiectomy were 1.75 days,225°,3.5 and 3.155 g/L;the area under the ROC curve(AUC)were 0.893,0.718,0.812 and 0.770,respectively.Conclusion The duration of onset,torsion angle,TWIST score,and FIB level are independent risk factors for testicular torsion patients with testicular resection as the outcome,and these indicators have certain predictive value.

Objective:To analyze the costs and effectiveness of five common screening modes and genetic screening for thalassemia in China in order to find the optimal way and provide evidence for the implementation of thalassemia prevention and control projects in Hunan Province.Methods:From June 2020 to April 2021, 12 971 couples from 14 cities and autonomous prefectures in Hunan Province were selected as the study population. The diagnosis of thalassemia was based on the results of genetic testing. Results of routine blood test and hemoglobin electrophoresis were collected and analyzed. The efficacy of five screening modes, at the cut-off value of <80 fl or 82 fl for the mean corpuscular volume (MCV), was analyzed by positive predictive value, negative predictive value, Jorden index and cost-effectiveness ratio. Sensitivity analysis was used to assess the feasibility of genetic screening at different costs after fixing the costs of routine blood and hemoglobin electrophoresis. The five thalassemia screening models are as follows: Mode 1: The woman had a blood routine test first. If the result was positive, the spouse required a blood routine test. If both results were positive, a thalassemia gene test should be offered to the couple. Mode 2: Both husband and wife were screened by blood routine and hemoglobin electrophoresis. If one or both of them were positive, both would be tested for thalassemia gene. Mode 3: The couple received blood routine tests initially. If either was positive, both should receive hemoglobin electrophoresis testing. If either was positive, both parties will conduct thalassemia gene testing. Mode 4: The woman was screened by blood routine and hemoglobin electrophoresis. If any one of them was positive, the woman would be tested for thalassemia gene. If the gene test result was positive, the spouse should receive thalassemia gene. Mode 5: Both spouses conducted a blood routine test. If either was positive, both would conduct hemoglobin electrophoresis test. If both were positive, both spouses should receive thalassemia gene testing. Gene testing mode: The woman would be tested for thalassemia, and her spouse would have thalassemia test too if her result was positive.Results:When using MCV<80 fl as the cut-off for diagnosing thalassemia, the Youden indices of the five prenatal screening modes in Hunan Province were 0.551, 0.639, 0.898, 0.555 and 0.356, while when using MCV<82 fl as the cut-off, the Youden indices were 0.549, 0.629, 0.851, 0.548 and 0.356. When the MCV cut-off value was <80 fl, the missed diagnosis rates of the five screening modes were 44.44%, 0.00, 0.00, 18.52% and 62.96%, and the cost-effectiveness ratios were 21 709, 250 939, 76 870, 138 463 and 92 860 yuan (RMB)/couple, respectively. When the price of genetic testing was lower than 55 yuan (RMB), the cost-effectiveness ratio of genetic screening was lower than that of Mode 3.Conclusions:MCV<80 fl can be considered as the positive criteria in blood routine screening for thalassemia in Hunan Province, and the cost-effectiveness ratio of Mode 3 (the couple received blood routine tests initially. If either was positive, both should receive hemoglobin electrophoresis testing. If either was positive, both parties will conduct thalassemia gene testing) is the best. Genetic screening has certain advantages with the decreasing price.

Objective : Angiotensin Ⅱ Type 2 receptor (AT2R) is a major receptor of angiotensin Ⅱ , which has a protective effect on damage to various tissues and organs．In this study，an overexpression plasmid of AT2R was con- structed to explore the effect of overexpression of AT2R on lipopolysaccharide ( LPS ) -induced inflammatory re- sponse in mouse hepatocytes (AML12 cells) ． Methods : Mice tissues and organs were used as samples to amplify the gene of interest (AT2R) fragments containing EcoR Ⅰ and Hind Ⅲ restriction sites，and then the final product was obtained by digestion and ligation．The final positive clones were sequenced and identified．The pCMV-Flag-N- AT2R plasmid was transfected into HEK 293T cells，and the expression of Flag protein was detected by the Western blot method after 24 h．AT2R expression on AML12 cells was observed by Western blot and laser confocal microsco- py．AML12 cells were treated differently and divided into control group，LPS treatment group，pCMV-Flag-N-AT2R group and pCMV-Flag-N-AT2R + LPS group，and cell viability was detected by CCK8．Western blot detected PCNA proteins and observed cell proliferation ; Cytoinflammatory factor levels were detected by qPCR ; Western blot detec- ted the expression level of nuclear transcription factor NF-кB (p65) in cells. Results : The identification and se- quencing results of EcoR I and Hind III double restriction showed that pCMV-Flag-N-AT2R plasmid was successful- ly constructed，and the detection results of the Western blot method showed successful expression of AT2R protein. Laser confocal observed that there were AT2R receptors on AML12 cells，and AT2R recombinant plasmids could be expressed on AML12; Compared with the control group，the viability and proliferation ability of LPS-treated AML12 cells were weakened，while the levels of IL-6 and TNF-α increased，and the expression level of nuclear transcription factor NF-кB (p65) increased．Compared with the LPS group，the viability and proliferation of cells in AML12 cells treated with pCMV-Flag-N-AT2R and LPS were enhanced，while the levels of IL-6 and TNF-α decreased．The ex- pression of the nuclear transcription factor NF-кB ( p65 ) decreased． Conclusion AT2R overexpression plasmids were successfully constructed and successfully expressed on AML12 cells，and AT2R could inhibit the inflammatory response of LPS-induced AML12 cells.

Objective To investigate the type and consumption of sanitary insecticides used in Putuo District of Shanghai， determine the current resistance of Aedes albopictus to the insecticides， and explore the causes of regional variations in insecticide resistance spectrum. Methods Public and private institutions of pest control operation were investigated on the use of sanitary insecticides. Dipping method and tube method were used to measure the insecticide resistance of Aedes albopictus， including larvae and adults. Results The main insecticides used in residential areas and governmental units was β-cypermethrin， while that in markets and public environment was propoxur. In addition， and the insecticides in dengue control program was λ-cyhalothrin. Aedes albopictus larvae had medium resistance to parathion， and were sensitive to propoxur， with insignificant change within three years. Their resistance to permethrin and deltamethrin was medium and high， respectively. Moreover， resistance to β-cypermethrin increased over years. In contrast， resistance of adult Aedes albopictus differed by area， except consistently being sensitive to fenitrothion. Conclusion Multiple sanitary insecticides have been used in Putuo District. In addition， Aedes albopictus has different resistance to these insecticides by area. It suggests that resistance surveillance should be promoted， which may be crucial for scientific application of insecticides and impede the development of potential resistance.

ObjectiveThis study aims to investigate the efficacy and underlying mechanism of Da Chaihutang （DCHT） in treating hepatocellular carcinoma （HCC） in vitro and in vivo. MethodWe employed methyl thiazolyl tetrazolium （MTT） assay and crystal violet staining to observe the proliferation of Hepa1-6 liver cancer cells treated with DCHT at different doses （0， 125， 250， 500， 1 000 mg·L^-1） for different time periods （1， 2， 4， 8 days）. The orthotopic liver cancer model was established by injection of 1×10⁶ Hepa1-6 cells into mouse， and then the model mice were randomly assigned into six groups： blank， model， DCHT （0.21， 0.625， 1.875 g·kg^-1， ig， qd）， and positive control （5-fluorouracil， 25 mg·kg^-1， ip， qod）. After 14 days of administration， the mice were sacrificed， and the liver samples were collected and fixed in 4% paraformaldehyde for hematoxylin-eosin （HE） staining. The Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform （TCMSP）， Cytoscape 3.7.2， STRING， and DAVID were used for the searching of the key targets of DCHT in treating HCC， the construction of protein-protein interaction （PPI） network， and the Kyoto Encyclopedia of Genes and Genomes （KEGG） pathway enrichment. Quantitative real-time PCR was performed to determine the mRNA level of interleukin-6 （IL-6） in Hepa1-6 cells and liver tissue. Western blotting was employed to measure the protein levels of the proteins involved in the mitogen-activated protein kinase （MAPK） and signal transducers and activators of transcription 3 （STAT3） signaling pathways. ResultDCHT （500， 1 000 mg·L^-1） treatment for 4 and 8 days inhibited the proliferation of Hepa1-6 cells in a dose- and time-dependent manner （P<0.05）. The in vivo assay showed that DCHT （high dose， 1.875 g·kg^-1） treatment for 14 days led to high differentiation and unobvious heterogeneity of HCC cells and small necrotic area compared with the model group. Network pharmacology analysis predicted that the potential targets of DCHT in the treatment of HCC were mainly the inflammation cytokines such as IL-6， interleukin-1β （IL-1β）， and tumor necrosis factor-alpha （TNF-α） in HCC microenvironment. The potential signaling pathways involved in the treatment were mainly associated with HCC growth and differentiation， including MAPK and STAT3 signaling pathways. Compared with the blank group， DCHT （1 000 mg·L^-1） treatment for 1， 2， 4， and 8 days down-regulated the mRNA level of IL-6 in Hepa1-6 cells （P<0.05）. Similar results were observed in the livers of mice treated with DCHT （0.625， 1.875 g·kg^-1）. The in vitro assay demonstrated that DCHT （1 000 mg·L^-1） treatment for 4 and 8 days and DCHT （500， 1 000 mg·L^-1） treatment inhibited the phosphorylation of extracellular signal-regulated kinases 1/2 （ERK1/2）， c-Jun NH₂-terminal kinase/stress-activated protein kinase （JNK）， p38 MAPK， and STAT3 in a dose- and time-dependent manner （P<0.05）. The in vivo assay showed that DCHT （0.625 and 1.875 g·kg^-1） treatment only inhibited the phosphorylation of p38 MAPK and STAT3 （P<0.05）. ConclusionThe present study indicates that DCHT can inhibit liver cancer cell proliferation by regulating p38 MAPK/IL-6/STAT3 signaling pathway.

Objective:To investigate the epidemiological characteristics of hospitalized children with respiratory syncytial virus (RSV)-associated acute lower respiratory tract infection (ALRTI), and to analyzed the risk factors for severe infection.Methods:The epidemiological and clinical data of hospitalized children with ALRTI and positive RSV test from Children′s Hospital of Fudan University from January 2013 to December 2018 were retrospectively analyzed.The hospitalized children from October 2016 to November 2017 were selected by random singular sequence and divided into severe infection group and non-severe infection group. The clinical data of the two groups were compared. Multivariate logistic regression analysis was used to analyze the risk factors of severe RSV-associated ALRTI.Results:A total of 34 192 hospitalized children were diagnosed with ALRTI, and 8 113(23.73%) children were positive for respiratory tract viruses, including 4 028(11.78%) children with RSV infection, which was higher than other common respiratory tract viruses. Among the 4 028 RSV-positive children, 2 550(63.31%) were under six months of age, 3 623(89.95%) were under two years of age. The detection rates of RSV in spring, summer, autumn and winter were 6.47%(553/8 551), 2.46%(176/7 161), 12.85%(1 042/8 111) and 21.77%(2 257/10 369), respectively. In 347 hospitalized children with RSV-associated ALRTI, 54 cases were severe cases. Multivariate logistic regression analysis showed that RSV-positive patients complicated with respiratory diseases ( Z=3.43), cardiovascular diseases ( Z=4.96), non-exclusive breast-feeding ( Z=-1.97) and premature birth ( Z=-1.98) were independent risk factors for severe RSV-associated ALRTI (all P<0.050). Conclusions:RSV is the most important and common viral pathogen in hospitalized children with ALRTI in Shanghai, and infants under six months of age are the most susceptible to RSV. RSV patients complicated with respiratory diseases, cardiovascular diseases, non-exclusive breast-feeding and premature birth are more likely to develope severe RSV-associated ALRTI.

OBJECTIVE: To assess the association of C677T polymorphism of methylenetetrahydrofolate reductase (MTHFR) gene with autistic behavior and inheritance pattern of children patients. METHODS: Ninety three autism patients were selected as the study group, whilst 93 healthy children were selected as the control group. The C677T genotype of the MTHFR gene was determined, and the correlation between the genotype and the autistic behavior and inheritance pattern were investigated. RESULTS: MTHFR gene C677T locus revealed three genotypes CC, CT and TT. Compared with the control group, the study group had fewer CC genotype but more TT genotype (P<0.05). Individuals with the three genotypes showed a statistically significant difference in the frequencies of four problem behaviors (P<0.05). Regression analysis showed that at least one T allele encoding the degree of 1 and 2 for the 4 problem behaviors that were statistically different. MTHFR gene C677T genotype was associated with autism under the recessive inheritance model and allelic inheritance model (P<0.05). CONCLUSION The C677T polymorphism of the MTHFR gene is associated with autistic behaviors. Children with the TT genotype or T allele are at higher risk of developing autism, particularly direct gaze, complex limb movements, self-injurious behavior and hyperactivity 1 and 2 related with the degree of coding.
Autistic Disorder/genetics* ; Child ; Genetic Predisposition to Disease ; Genotype ; Humans ; Inheritance Patterns ; Methylenetetrahydrofolate Reductase (NADPH2)/genetics* ; Problem Behavior

Tumor-associated macrophages (TAMs), one of the dominating constituents of tumor microenvironment, are important contributors to cancer progression and treatment resistance. Therefore, regulation of TAMs polarization from M2 phenotype towards M1 phenotype has emerged as a new strategy for tumor immunotherapy. Herein, we successfully initiated antitumor immunotherapy by inhibiting TAMs M2 polarization via autophagy intervention with polyethylene glycol-conjugated gold nanoparticles (PEG-AuNPs). PEG-AuNPs suppressed TAMs M2 polarization in both in vitro and in vivo models, elicited antitumor immunotherapy and inhibited subcutaneous tumor growth in mice. As demonstrated by the mRFP-GFP-LC3 assay and analyzing the autophagy-related proteins (LC3, beclin1 and P62), PEG-AuNPs induced autophagic flux inhibition in TAMs, which is attributed to the PEG-AuNPs induced lysosome alkalization and membrane permeabilization. Besides, TAMs were prone to polarize towards M2 phenotype following autophagy activation, whereas inhibition of autophagic flux could reduce the M2 polarization of TAMs. Our results revealed a mechanism underlying PEG-AuNPs induced antitumor immunotherapy, where PEG-AuNPs reduce TAMs M2 polarization via induction of lysosome dysfunction and autophagic flux inhibition. This study elucidated the biological effects of nanomaterials on TAMs polarization and provided insight into harnessing the intrinsic immunomodulation capacity of nanomaterials for effective cancer treatment.