Objective To establish an indirect enzyme-linked immunosorbent assay(ELISA)method for testing the concentration of a monoclonal antibody target tumor necrosis factor-α(TNF-α)in animal serum.Methods The critical parameters of the method including coating concentration of human TNF-α,source,concentration and stability of HRP-labeled goat anti-human immunoglobulin G(IgG)were investigated.The specificity,accuracy,precision,linearity and Limited of Determination of the method were investigated.Results The critical parameters of the method were confirmed as below:TNF-α was coated at 400 ng·mL-1;HRP labeled goat anti-human IgG antibody was diluted at 1:3.0 ×105;the diluted horseradish peroxidase labeled goat anti-human IgG antibody is well stored at 4 ℃ for 3 days.Meanwhile the method was confirmed to have good specificity,the recovery rate ranged from 84.00％to 106.82％,the coefficient of variation of different antibody concentration levels were no more than 10％;the method had a good linearity and the standard curve was y=(-8.37×103-2.37 × 106)/[1+(x/29.80)106]+2.37 × 106(R2=0.999);the limit of quantification was 1 ng·mL-1,all of which met the requirements.Conclusion A accurate and robust ELISA method was developed to test the concentration of anti-TNF-α monoclonal antibody in serum.

Chlamydia psittaci is a zoonotic respiratory infection.To provide a reference basis for psittaciosis-specific pre-vention and control,we conducted a retrospective analysis reviewing the clinical and epidemiological characteristics of three pa-tients.All patients had a history of avian or bird contact,and their clinical symptoms were primarily high fever and pneumonia.Chlamydia psittaci nucleic acid testing was conducted on the patients'close contacts,surrounding occupational populations,and their living environments.No positive findings were observed among the cases'close contacts and surrounding occupational populations,whereas environmental positivity was detected in the feces of the patients'neighbors(25.0％,3/12)and in the ef-fluent(22.2％,2/9)of a wild bird activity area.Positive samples were subjected to ompA gene sequencing,which revealed that the strains in three patients were genetically sequence-identical to that found in pigeon feces.This study enhances under-standing of the epidemic caused by Chlamydia psittaci,elucidates its transmission mechanisms,and provides critical insights for subsequent investigations and management of the disease.

This study was aimed at surveying the distribution of Himalayan marmots in Gaotai County,Zhangye City,Gan-su Province,and analyzing the risk of plague occurrence,to provide a scientific basis for plague prevention and control in the area.On the basis of preliminary evidence,we used a combined approach of on-site investigation and Global Positioning System(GPS)positioning to survey the distribution and natural geographical landscape characteristics of the Himalayan marmot in Gaotai County.Population information and customs in the marmot distribution area were investigated through methods such as data retrieval and household visits.Himalayan marmots were captured through bow-shaped trapping,and blood,liver,spleen,heart,lung,kidney,femur,and surface-intermediate host samples were collected.Pathological and serological tests were conducted according to the Plague Diagnosis Criteria(WS279-2008).In Xinba Town,Gaotai County,Himalayan marmots were found in 16 villages(53.33％).Heping Village had the largest distribution area(100％),and was followed by Zhaoyi Village(79.17％)and Xishang Village(55.32％),whereas the other areas had values below 50％.Regarding the direct threats posed by marmots,Heping Village,Dongda Vil-lage,and Zhaoyi Village had values of 100％,Xishang Village had a value of 90.59％,and the other areas had values below 50％.The flea infestation rate on marmots'bodies was 59.62％,and the flea index was 1.48％(77/52).The main surface-in-termediate hosts were steppe ticks,body lice,Xenopsylla siberica,and Nosopsyllus laeviceps,accounting for 72.31％(517/715),16.92％(121/715),6.43％(46/715),and 4.20％(30/715),respectively,with steppe ticks being the dominant species.A total of 52 liver and spleen tissue samples from marmots,and 715 surface-intermediate hosts samples,were collected for bacterial culture.No Yersinia pestis was found.RIHA testing for plague F1 antigen was negative in all samples.Additionally,52 blood serum samples from marmots were collected,and IHA testing for plague-specific F1 antibodies was negative in all samples.The natural plague foci of marmots in the Qilian Mountains and Altyn Mountains is affected by factors including con-tinuing increases in temperature and grassland degradation.As a result,the marmot plague hosts continually migrate to sur-rounding non-epidemic areas,particularly in Gaotai County,Zhangye City.Local residents often hunt wild rabbits and mar-mots,thus posing a risk of human plague outbreaks if animal plague occurs.This county is recommended to be included in the province's plague surveillance and epidemic source investigation.Additionally,efforts should be strengthened in health educa-tion,medical personnel training,and management of key individuals and emergency reserves.If necessary,protective measures should be taken in marmot and flea control,to prevent the occurrence and spread of plague.

Chlamydia psittaci is a zoonotic respiratory infection.To provide a reference basis for psittaciosis-specific pre-vention and control,we conducted a retrospective analysis reviewing the clinical and epidemiological characteristics of three pa-tients.All patients had a history of avian or bird contact,and their clinical symptoms were primarily high fever and pneumonia.Chlamydia psittaci nucleic acid testing was conducted on the patients'close contacts,surrounding occupational populations,and their living environments.No positive findings were observed among the cases'close contacts and surrounding occupational populations,whereas environmental positivity was detected in the feces of the patients'neighbors(25.0％,3/12)and in the ef-fluent(22.2％,2/9)of a wild bird activity area.Positive samples were subjected to ompA gene sequencing,which revealed that the strains in three patients were genetically sequence-identical to that found in pigeon feces.This study enhances under-standing of the epidemic caused by Chlamydia psittaci,elucidates its transmission mechanisms,and provides critical insights for subsequent investigations and management of the disease.

This study was aimed at surveying the distribution of Himalayan marmots in Gaotai County,Zhangye City,Gan-su Province,and analyzing the risk of plague occurrence,to provide a scientific basis for plague prevention and control in the area.On the basis of preliminary evidence,we used a combined approach of on-site investigation and Global Positioning System(GPS)positioning to survey the distribution and natural geographical landscape characteristics of the Himalayan marmot in Gaotai County.Population information and customs in the marmot distribution area were investigated through methods such as data retrieval and household visits.Himalayan marmots were captured through bow-shaped trapping,and blood,liver,spleen,heart,lung,kidney,femur,and surface-intermediate host samples were collected.Pathological and serological tests were conducted according to the Plague Diagnosis Criteria(WS279-2008).In Xinba Town,Gaotai County,Himalayan marmots were found in 16 villages(53.33％).Heping Village had the largest distribution area(100％),and was followed by Zhaoyi Village(79.17％)and Xishang Village(55.32％),whereas the other areas had values below 50％.Regarding the direct threats posed by marmots,Heping Village,Dongda Vil-lage,and Zhaoyi Village had values of 100％,Xishang Village had a value of 90.59％,and the other areas had values below 50％.The flea infestation rate on marmots'bodies was 59.62％,and the flea index was 1.48％(77/52).The main surface-in-termediate hosts were steppe ticks,body lice,Xenopsylla siberica,and Nosopsyllus laeviceps,accounting for 72.31％(517/715),16.92％(121/715),6.43％(46/715),and 4.20％(30/715),respectively,with steppe ticks being the dominant species.A total of 52 liver and spleen tissue samples from marmots,and 715 surface-intermediate hosts samples,were collected for bacterial culture.No Yersinia pestis was found.RIHA testing for plague F1 antigen was negative in all samples.Additionally,52 blood serum samples from marmots were collected,and IHA testing for plague-specific F1 antibodies was negative in all samples.The natural plague foci of marmots in the Qilian Mountains and Altyn Mountains is affected by factors including con-tinuing increases in temperature and grassland degradation.As a result,the marmot plague hosts continually migrate to sur-rounding non-epidemic areas,particularly in Gaotai County,Zhangye City.Local residents often hunt wild rabbits and mar-mots,thus posing a risk of human plague outbreaks if animal plague occurs.This county is recommended to be included in the province's plague surveillance and epidemic source investigation.Additionally,efforts should be strengthened in health educa-tion,medical personnel training,and management of key individuals and emergency reserves.If necessary,protective measures should be taken in marmot and flea control,to prevent the occurrence and spread of plague.

Objective: To analyze the clinical phenotype and screen the genetic mutations of hereditary deafness in three deaf families to clarify their molecular biology etiology. Methods: From January 2019 to January 2020, three deaf children and family members were collected for medical history, physical examination, audiology evaluation, electrocardiogram and cardiac color Doppler ultrasound, temporal bone CT examination, and peripheral blood DNA was obtained for high-throughput sequencing of deafness genes. Sanger sequencing was performed to verify the variant sites among family members. The pathogenicity of the variants was evaluated according to the American College of Medical Genetics and Genomics. Results: The probands in the three families had deafness phenotypes. In family 1, proband had multiple lentigines, special facial features, growth retardation, pectus carinatum, abnormal skin elasticity, cryptorchidism and other manifestations. In family 2, proband had special facial features, growth retardation and abnormal heart, and the proband in family 3 had growth retardation and abnormal electrocardiogram. Genetic testing of three families detected three heterozygous mutations in the PTPN11 gene: c.1391G>C (p.Gly464Ala), c.1510A>G (p.Met504Val), c.1502G>A (p.Arg501Lys). All three sites were missense mutations, and the mutation sites were highly conserved among multiple homologous species. Based on clinical manifestations and genetic test results, proband 1 was diagnosed with multiple lentigines Noonan syndrome, and probands 2 and 3 were diagnosed with Noonan syndrome. Conclusion: Missense mutations in the PTPN11 gene may be the cause of the disease in the three deaf families. This study enriches the clinical phenotype and mutation spectrum of the PTPN11 gene in the Chinese population.
Deafness/genetics* ; Genetic Testing ; Hearing Loss/genetics* ; Humans ; Male ; Mutation ; Phenotype ; Protein Tyrosine Phosphatase, Non-Receptor Type 11/genetics*

Objective:To explore the clinical efficacy of myofascial trigger point electric stimulation based on mirror therapy on phantom limb pain after lower limb amputation. Methods:From May to November, 2020, 50 patients with phantom limb pain after lower limb amputation were randomly divided into control group (n = 25) and experiment group (n = 25). Both groups accepted mirror therapy, while the experiment group received myofascial trigger point electric stimulation before mirror therapy, for four weeks. They were assessed with short-form of McGill Pain Questionnaire (SF-MPQ), Pittsburgh Sleep Quality Index (PSQI), Hamilton Anxiety Scale (HAMA), Timed 'Up & Go' Test (TUGT) and 6-minute walk test (6MWT) before and after treatment. Results:All the indexes improved in both groups after treatment (|t| > 8.210, P < 0.001), and improved more in the experiment group than in the control group (|t| > 5.103, P < 0.001), except the present pain intensity of SF-MPQ. Conclusion:Mirror therapy is effective on phantom limb pain after lower limb amputation in terms of pain, sleep, anxiety and walking, and the effect could be stronger after myofascial trigger point electric stimulation.

BACKGROUND: Zinc finger and BTB domain-containing protein 46 (Zbtb46) is a transcription factor identified in classical dendritic cells, and maintains dendritic cell quiescence in a steady state. Zbtb46 has been reported to be a negative indicator of acute myeloid leukemia (AML). We found that Zbtb46 was expressed at a relatively higher level in hematopoietic stem and progenitor cells (HSPCs) compared to mature cells, and higher in AML cells compared to normal bone marrow (BM) cells. However, the role of Zbtb46 in HSPCs and AML cells remains unclear. Therefore, we sought to elucidate the effect of Zbtb46 in normal hematopoiesis and AML cells. METHODS: We generated Zbtb46 and Zbtb46Mx1-Cre mice. The deletion of Zbtb46 in Zbtb46Mx1-Cre mice was induced by intraperitoneal injection of double-stranded poly (I). poly (C) (poly(I:C)), and referred as Zbtb46 cKO. After confirming the deletion of Zbtb46, the frequency and numbers of HSPCs and mature blood cells were analyzed by flow cytometry. Serial intraperitoneal injection of 5-fluorouracil was administrated to determine the repopulation ability of HSCs from Zbtb46 and Zbtb46 cKO mice. The correlation between Zbtb46 expression and prognosis was analyzed using the data from the Cancer Genome Atlas. To investigate the role of Zbtb46 in AML cells, we knocked down the expression of Zbtb46 in THP-1 cells using lentiviral vectors expressing small hairpin RNAs targeting Zbtb46. Cell proliferation rate was determined by cell count assay. Cell apoptosis and bromodeoxyuridine incorporation were determined by flow cytometry. RESULTS: The percentages and absolute numbers of HSPCs and mature blood cells were comparable in Zbtb46 cKO mice and its Zbtb46 littermates (Zbtb46vs. Zbtb46 cKO, HPC: 801,310 ± 84,282 vs. 907,202 ± 97,403, t = 0.82, P = 0.46; LSK: 86,895 ± 7802 vs. 102,210 ± 5025, t = 1.65, P = 0.17; HSC: 19,753 ± 3116 vs. 17,608 ± 3508, t = 0.46, P = 0.67). The repopulation ability of HSCs from Zbtb46Mx1-Cre mice was similar to those from Zbtb46 control (P = 0.26). Zbtb46 had elevated expression in AML cells compared to total BM cells from normal control. Knockdown of Zbtb46 in THP-1 cells led to a significant increase in cell apoptosis and reduced cell growth and proliferation. CONCLUSION Collectively, our data indicate that Zbtb46 is essential for survival and proliferation of AML cells, but dispensable for normal hematopoiesis.

Objective To investigate the protective effects of Salvianolic acid on myocardial ischemic injury of rats from the aspect of inhibiting inflammatory reaction.Methods Sixty Sprague Dawley (SD) male rats were randomly divided into four groups (n =15per group):sham operation group,model group,experimental-high (36 mg · kg-1) group,experimental-low (18mg · kg-1) group with intraperitoneal injection dosing.Same amount of distilled water was injected to sham operation group and model group,lasting for 4 days.Myocardial ischemia model was established by coronary artery ligation of left anterior descending (LAD) branch after the last administration.Six hours after the operation,the blood of eye canthus was collected to assay the levels of creatine kinase(CK),MB isoenzyme of creatine kinase (CK-MB) and lactate dehydrogenase (LDH) in serum.The heart was collected 24 h after the operation,stained by hematoxylin eosin (HE)and checked the pathological change of the cardiac muscle tissue,assayed the cardiac troponin (cTnT),tumor necrosis factor α (TNF-α) in serum and myeloperoxidase (MPO) was measured in myocardial tissue homogenate.Results After 6 hours' myocardial ischemia,the serum CK in sham operation group was (529.61 ± 141.93) U · L-1,CK-MB was (708.12 ±385.93) U · L-1,LDH was (330.12 ±158.38) U · L-1 and cTnT was (294.12 ±55.10)pg· mL-1 after 24 hours' myocardial ischemia.The serum CK in model group was (996.42 ± 413.42) U · L-1,CK-MB was (1346.11 ±558.30) U · L-1,LDH was (520.12 ±154.76) U · L-1and cTnT was (513.00 ±69.71)pg · mL-1after 24 hours' myocardial ischemia.The serum CK in experimental-low and experimental-high groups were (499.45 ±159.33),(514.91 ±98.82)U · L-1,CK-MB were (831.42 ±385.11),(592.10 ±206.32)U·L-1,LDH were (462.62 ±229.68),(437.72 ± 175.80)U· L-1,and cTnT were (431.12 ±106.00),(338.80 ± 76.92)pg · mL-1after 24 hours' myocardial ischemia.Compared with the sham operation group,the CK,CK-MB and LDH value in the serum of model group increased significantly (P ＜ 0.05).Compared with the model group,the salvianolic acid significantly both can decrease the CK and CK-MB in serum (P ＜ 0.05).Compared with sham operation group,the cTnT value in the serum of model group increased significantly (P ＜ 0.05).Compared with model group,high dosage of salvianolic acid could significantly decrease the cTnT value (P ＜ 0.05).After 24 hours'myocardial ischemia,the TNF-α in serum of sham operation group was (118.90 ± 17.58) pg · mL-1,the MPO in heart tissue was (32.25 ± 3.75) U · L-1.The TNF-α in serum of model group was (156.00 ± 28.24) pg · mL-1,the MPO in heart tissue was (104.83 ±22.87) U · L-1.The TNF-α in serum of experimental-low and experimental-high groups were (136.05 ± 34.00),(125.06 ± 30.13) pg · mL-1,the MPO in heart tissue were (91.70 ±21.57),(68.00 ± 18.47) U · L-1.Compared with sham operation group,the TNF-α values in the serum and the MPO value in model group increased significantly (P ＜ 0.05).Compared with model group,these values of experimental-high group decreased with significantly (all P ＜ 0.05).Conclusion Salvianolic acid showed the protection of myocardial tissue,which also could significantly reduce the myocardial cell injury and inflammatory cell infiltration of cardiac myocytes,which could be related with the decreased levels of TNF-α and MPO.