Objective To clone and express the heat shock cognate protein 20 (SjHsc20) of Schistosoma japonicum, and to preliminarily investigate its biological characteristics. Methods The target fragment of the SjHsc20 gene was amplified using PCR assay and cloned into the pET-28a(+) expression plasmid to generate the recombinant expression vector pET-28a(+)-SjH-sc20, which was then transformed into Escherichia coli BL21 (DE3) competent cells. The recombinant SjHsc20 (rSjHsc20) protein was induced with isopropyl β-D-thiogalactopyranoside (IPTG) and purified, and the expression of the rSjHsc20 protein was checked with sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The immunogenicity of the rSjHsc20 protein was detected using Western blotting, and the transcriptional levels of SjHsc20 were quantified in S. japonicum worms at different developmental stages and in male and female adult worms using real-time quantitative PCR (RT-qPCR) assay. Thirty female BALB/c mice at ages 6 to 8 weeks were divided into three groups, including the rSjHsc20 immunization group, the PBS control group, and the ISA 206 adjuvant group, of 10 mice in each group. Mice in the rSjHsc20 immunization group were subcutaneously immunized with 20 μg rSjHsc20 on days 1, 15 and 31, and animals in the PBS control group were subcutaneously injected with the same volume of PBS on days 1, 15 and 31, while mice in the ISA 206 adjuvant group were subcutaneously immunized with the same volume of ISA 206 adjuvant on days 1, 15 and 31, respectively. All mice in each group were infected with (40 ± 2) S. japonicum cercariae via the abdomen 14 day following the last immunization. Levels of serum specific IgG and its subtypes IgG1 and IgG2 antibodies against rSjHsc20, and the serum titers of anti-rSjHsc20 antibody were detected in mice using indirect enzyme-linked immunosorbent assay (ELISA). All mice were sacrifice 42 days post-infection, and S. japonicum worms were collected from the hepatic portal vein and counted. The eggs per gram (EPG), worm burden reductions and egg burden reductions were estimated to evaluate the protective efficacy of the rSjHsc20 protein. Results The SjHsc20 gene had an open reading frame (ORF) with 756 bp in length and encoded 252 amino acids, and the rSjHsc20 protein had a relative molecular mass of approximately 29 kDa. The rSjHsc20 protein was recognized by the serum of mice infected with S. japonicum and the serum of mice immunized with the rSjHsc20 protein, indicating that rSjHsc20 had a good immunogenicity. There was a significant difference in the transcriptional levels of the SjHsc20 gene among the 7-day (1.001 4 ± 0.065 7), 12-day (2.268 3 ± 0.129 2), 21-day (1.378 5 ± 0.160 4), 28-day (1.196 4 ± 0.244 0), 35-day (1.646 3 ± 0.226 1), 42-day worms of S. japonicum (1.758 0 ± 0.611 1) (F = 38.45, P < 0.000 1), and the transcriptional level of the SjHsc20 gene was higher in the 12-day worms than in worms at other developmental stages (all P values < 0.000 1). The serum levels of anti-rSjHsc20 IgG antibody were 0.106 6 ± 0.010 7, 0.108 3 ± 0.010 4, and 0.553 2 ± 0.069 1 in the PBS control group, ISA 206 adjuvant group, and rSjHsc20 immunization group following the last immunization, respectively, and the serum levels of IgG1 antibody were 0.137 3 ± 0.054 0, 0.181 1 ± 0.096 8, and 1.765 8 ± 0.221 1, while the levels of IgG2a antibody were 0.280 3 ± 0.197 6, 0.274 0 ± 0.146 3, and 1.560 4 ± 0.106 0, respectively. There were significant differences in the serum levels of anti-rSjHsc20 IgG (F = 397.70, P < 0.000 1), IgG1 (F = 401.00, P < 0.000 1) and IgG2a antibodies (F = 229.70, P < 0.000 1) among the three groups, and the serum levels of anti-rSjHsc20 IgG, IgG1 and IgG2a antibodies were higher in the rSjHsc20 immunization group than in the PBS control group and the ISA 206 adjuvant group (all P values < 0.000 1). There was a significant difference in the IgG1/IgG2a ratio among the rSjHsc20 immunization group (1.177 2 ± 0.143 6), the PBS control group (0.428 4 ± 0.199 8) and the ISA 206 adjuvant group (0.559 9 ± 0.181 1) (F = 43.97, P < 0.000 1), and the IgG1/IgG2a ratio was > 1 in the rSjHsc20 immunization group, which was higher than in the PBS control group and the ISA 206 adjuvant group (both P values < 0.000 1). The titers of serum anti-rSjHsc20 antibody were all above 1∶16 384 in the rSjHsc20 immunization group following immunizations on days 1, 15 and 31, indicating that the rSjHsc20 protein had a strong immunogenicity. The mean worm burdens were (16.60±5.75), (15.80±5.58) worms per mouse and (14.40±5.75) worms per mouse in the PBS control group, the ISA 206 adjuvant group and the rSjHsc20 immunization group 42 days post-infection with S. japonicum cercariae (F = 0.50, P > 0.05), and the EPG were 68 370 ± 22 690, 67 972 ± 19 502, and 41 075 ± 13 251 in the PBS control group, the ISA 206 adjuvant group and the rSjHsc20 immunization group (F = 4.55, P < 0.05), with lower EPG in the PBS control group and the ISA 206 adjuvant group than in the rSjHsc20 immunization group (both P values < 0.05). Immunization with the rSjHsc20 protein resulted in a worm burden reduction of 13.25% and an egg burden reduction of 39.92% relative to the PBS control group. Conclusions SjHsc20 is successfully cloned and expressed, and the rSjHsc20 protein induces partial immunoprotective effects in mice, which provides a basis for deciphering the biological functions of SjHsc20 and assessing the potential of SjH-sc20 as a vaccine candidate.

Debates persist regarding the efficacy and safety of azvudine, particularly its real-world outcomes. This study involved patients aged ≥60 years who were admitted to 25 hospitals in mainland China with confirmed SARS-CoV-2 infection between December 1, 2022, and February 28, 2023. Efficacy outcomes were all-cause mortality during hospitalization, the proportion of patients discharged with recovery, time to nucleic acid-negative conversion (T _NANC), time to symptom improvement (T _SI), and time of hospital stay (T _HS). Safety was also assessed. Among the 5884 participants identified, 1999 received azvudine, and 1999 matched controls were included after exclusion and propensity score matching. Azvudine recipients exhibited lower all-cause mortality compared with controls in the overall population (13.3% vs. 17.1%, RR, 0.78; 95% CI, 0.67-0.90; P = 0.001) and in the severe subgroup (25.7% vs. 33.7%; RR, 0.76; 95% CI, 0.66-0.88; P < 0.001). A higher proportion of patients discharged with recovery, and a shorter T _NANC were associated with azvudine recipients, especially in the severe subgroup. The incidence of adverse events in azvudine recipients was comparable to that in the control group (2.3% vs. 1.7%, P = 0.170). In conclusion, azvudine showed efficacy and safety in older patients hospitalized with COVID-19 during the SARS-CoV-2 omicron wave in China.

This research study focuses on addressing the limitations of current neuropathic pain (NP) treatments by developing a novel dual-target modulator, E0199, targeting both Na_V1.7, Na_V1.8, and Na_V1.9 and K_V7 channels, a crucial regulator in controlling NP symptoms. The objective of the study was to synthesize a compound capable of modulating these channels to alleviate NP. Through an experimental design involving both in vitro and in vivo methods, E0199 was tested for its efficacy on ion channels and its therapeutic potential in a chronic constriction injury (CCI) mouse model. The results demonstrated that E0199 significantly inhibited Na_V1.7, Na_V1.8, and Na_V1.9 channels with a particularly low half maximal inhibitory concentration (IC₅₀) for Na_V1.9 by promoting sodium channel inactivation, and also effectively increased K_V7.2/7.3, K_V7.2, and K_V7.5 channels, excluding K_V7.1 by promoting potassium channel activation. This dual action significantly reduced the excitability of dorsal root ganglion neurons and alleviated pain hypersensitivity in mice at low doses, indicating a potent analgesic effect without affecting heart and skeletal muscle ion channels critically. The safety of E0199 was supported by neurobehavioral evaluations. Conclusively, E0199 represents a ground-breaking approach in NP treatment, showcasing the potential of dual-target small-molecule compounds in providing a more effective and safe therapeutic option for NP. This study introduces a promising direction for the future development of NP therapeutics.

Objective To develop a multiplex PCR assay for simultaneous detection of four intestinal parasites, including Giardia duodenalis, Cryptosporidium parvum, Enterocytozoon bieneusi and Moniezia, and to preliminarily evaluate its detection efficiency. Methods Four pairs of specific primers were designed based on the conserved sequences of the corresponding genes of G. duodenalis (GenBank accession number: XM_001710026.2), C. parvum (GenBank accession number: XM_626998.1), E. bieneusi (GenBank accession number: KJ719492.1) and Moniezia (GenBank accession number: OM296991.1) retrieved from the GenBank database, and a multiplex PCR assay for simultaneous detection of G. duodenalis, C. parvum, E. bieneusi and Moniezia was developed and optimized. A total of 116 fresh goat stool samples were collected from four goat farms in Zhanjiang City, Guangdong Province during the period from October to December 2022, including 96 samples used for evaluating the detection efficacy of the multiplex PCR assay, and 20 samples as baseline controls for sample testing. Genomic DNA extracted from 96 goat stool samples was tested using the single-target PCR assay and the developed multiplex PCR assay, and the sensitivity, specificity, positive predictive value, and negative predictive value of the multiplex PCR assay were evaluated for detection of G. duodenalis, C. parvum, E. bieneusi and Moniezia DNA in goat stool samples with the single-target PCR assay as the gold standard. Results The multiplex PCR assay developed in this study allowed simultaneous amplification of specific gene fragments of G. duodenalis, C. parvum, E. bieneusi and Moniezia, with 1 400, 755, 314 bp and 585 bp in sizes, respectively, and the detection limit was 10² and higher copies of parasite DNA clones, while the multiplex PCR assay was negative for gene amplification of Schistosoma japonicum, Fasciola hepatica, Echinococcus granulosus, Blastocystis hominis and Homalogaster paloniae. Single-target PCR assay and the developed multiplex PCR assay were employed to test DNA samples extracted from 96 goat stool samples, and single-target PCR assay tested positive in 40 goat stool samples (41.67%), including 39 positive samples tested with the multiplex PCR assay, with a mean coincidence rate of 97.50% (39/40). The multiplex PCR assay tested positive for G. duodenalis DNA in 26 goat stool samples (27.10%), C. parvum DNA in 22 samples (22.90%), E. bieneusi DNA in 24 samples (25.00%), and Moniezia in 9 samples (9.40%), which was consistent with the detection using the single-target PCR assay. The sensitivity, negative predictive value, and positive predictive value of the multiplex PCR assay were 96.15%, 95.83%, 100.00% and 100.00%, 98.90%, 98.92%, 100.00% and 100.00%, 100.00%, 100.00%, 100.00% and 100.00% for detection of G. duodenalis, C. parvum, E. bieneusi and Moniezia DNA in goat stool samples, respectively, if the single-target PCR assay served as the gold standard. Conclusion A highly sensitive and specific multiplex PCR assay has been developed for simultaneous detection of G. duodenalis, C. parvum, E. bieneusi and Moniezia in goats, which is suitable for rapid, large-scale screening of intestinal parasites in sheep stool samples.

Caprine enterovirus(CEV)is one of the most important infectious diarrheal diseases for goats.In order to understand the prevalence of CEV in different areas of Jiangsu province,410 goat fecal samples(212 diarrhea and 198 non-diarrhea samples)were collected from goat farms in 8 re-gions.127 CEV positive samples were detected by RT-PCR,and the positive rates varied greatly from 12.50％to 62.50％among different areas,with a total positive rate as 30.98％.Through the significance analysis of the positive rate of diarrhea and non-diarrhea samples,CEV was found to be one of the important pathogens causing diarrhea in goats.The results of homology and genetic evolution analysis for the 5'-UTR sequences showed that the EV-G5、G7、G21、G22 and G23 were epidemic types in Jiangsu province,and G21-G23 are new found CEV types.The homologies of 5'-UTR,VP1 and 2AB genes with the classical CEV strains in China were 78.6％-95.8％,56.9％-95.0％and 77.9％-98.4％,respectively.Of these sequences,the 5'-UTR and 2AB gene of HaiMen-SJH strain was significantly different from other sequences,and belonged to an independent branch in the genetic evolution trees.According to the results of epidemiological survey,the infection of CEV was widespread in most areas of Jiangsu province,and the epidemic types and situation varied in different areas.This study will enrich the epidemiological data of CEV in China,and provide the-oretical basis for the targeted prevention and control of new enterovirus infections in sheep.

Objective:To discuss their effects of PLA(polylactic acid)/PTMC(polytrimethylene carbonate)and PLA/PTMC-calcium phosphate[Ca3(PO4)2]composite porous scaffolds prepared by 3D printing technology on bone marrow mesenchymal stem cells(BMSCs)of the rabbits,and to clarify their application values in bone defect repairment.Methods:After mixing the materials,PLA/PTMC and PLA/PTMC-Ca3(PO4)2 filaments were prepared by desktop filament extruder.The scaffolds were designed by CATIA V5-6R2019 modeling software and fabricated using CreatBot F430 3D printer.The chemical structure of the PLA/PTMC-Ca3(PO4)2 scaffold was detected by infrared spectroscopy.In vitro degradation experiments were used to detect the degradation weight loss rates and pH values of the two scaffolds.A contact angle measuring instrument was used to detect the hydrophilicities of the two scaffolds.The BMSCs were extracted from three newborn New Zealand white rabbits(2-5-day-old);CCK-8 method was used to detect the proliferation activities of the cells co-cultured with two scaffolds,and Alizarin red staining was used to observe the osteogenic differentiation of the cells co-cultured with two scaffolds.Results:Infrared spectroscopy confirmed the successful preparation of composite scaffolds containing PLA,PTMC,and β-Ca3(PO4)2.During degradation for 6-14 weeks,compared with PLA/PTMC scaffold,the degradation rates of the PLA/PTMC-Ca3(PO4)2 scaffold in lipase solution and phosphate-buffered saline(PBS)were significantly increased(P<0.05 or P<0.01).During degradation for 8-14 weeks,compared with PLA/PTMC scaffold,the pH value of the PLA/PTMC-Ca3(PO4)2scaffold in lipase solution was significantly increased(P<0.01).Compared with PLA/PTMC scaffold,the contact angle of the PLA/PTMC-Ca3(PO4)2 scaffold was significantly decreased(P<0.01).On days 5 and 7 of cell co-culture,compared with PLA/PTMC scaffold,the proliferation activity of the cells co-cultured with PLA/PTMC-Ca3(PO4)2 scaffold was significantly increased(P<0.05 or P<0.01).After 21 d of co-culture,both scaffolds overlapped with BMSCs and locally formed calcified nodules,which were stained orange by Alizarin red.Compared with PLA/PTMC scaffold,the number of mineralized calcium nodules in the cells co-cultured with PLA/PTMC-Ca3(PO4)2 scaffold was increased,with greater density and deeper color.Conclusion:The PLA/PTMC-Ca3(PO4)2 composite porous scaffolds containing PLA,PTMC,and β-Ca3(PO4)2 are successfully prepared by 3D printing technology.These scaffolds exhibit good degradation properties and show advantages in biocompatibility,hydrophilicity,and osteogenic induction;they are excellent materials for the bone defect repairment.

Ommaya reservoir implantation is generally used in the treatment of hydrocephalus and intraventricular drug administration. Ommaya reservoir implantation in the subarachnoid space of the spinal cord for the intrathecal drug administration has not been carried out in China, and only several reports can be retrieved from PubMed. About 60%-90% of untreated patients with spinal muscular atrophy type 2 (SMA2) who survive to adulthood often have complex scoliosis and joint deformities. Nusinersen is an effective drug for the treatment of SMA2. And the route of administration is intrathecal injection, which is difficult for patients with severe scoliosis. This article summarizes the process of Ommaya reservoir implantation and postoperative drug administration in a patient with complex scoliosis type SMA2, which provides a new method for clinical treatment of this disease.

In response to the problem that the traditional lower limb rehabilitation scale assessment method is time-consuming and difficult to use in exoskeleton rehabilitation training, this paper proposes a quantitative assessment method for lower limb walking ability based on lower limb exoskeleton robot training with multimodal synergistic information fusion. The method significantly improves the efficiency and reliability of the rehabilitation assessment process by introducing quantitative synergistic indicators fusing electrophysiological and kinematic level information. First, electromyographic and kinematic data of the lower extremity were collected from subjects trained to walk wearing an exoskeleton. Then, based on muscle synergy theory, a synergistic quantification algorithm was used to construct synergistic index features of electromyography and kinematics. Finally, the electrophysiological and kinematic level information was fused to build a modal feature fusion model and output the lower limb motor function score. The experimental results showed that the correlation coefficients of the constructed synergistic features of electromyography and kinematics with the clinical scale were 0.799 and 0.825, respectively. The results of the fused synergistic features in the K-nearest neighbor (KNN) model yielded higher correlation coefficients ( r = 0.921, P < 0.01). This method can modify the rehabilitation training mode of the exoskeleton robot according to the assessment results, which provides a basis for the synchronized assessment-training mode of "human in the loop" and provides a potential method for remote rehabilitation training and assessment of the lower extremity.
Humans ; Exoskeleton Device ; Reproducibility of Results ; Walking/physiology* ; Lower Extremity ; Algorithms ; Stroke Rehabilitation/methods*

Objective:To explore the safety and efficacy of laparoscopic non-blocking partial nephrectomy assisted by high power lateral green laser in the treatment of T 1a renal tumor. Methods:The clinical data of 10 patients with T1a stage renal tumor from February 2021 to April 2021 in department of urology, Gongli hospital affiliated to Naval Military Medical University were retrospectively analyzed. There were 7 males and 3 females, aged 47.0-74.0 years, with average of(58.8±9.7)years old. The diameter of the tumor ranged from 2.0 cm to 3.8 cm, with an average of (3.1±0.6)cm. There were 6 cases on the left side and 4 cases on the right side, locate on lumbar side in 9 cases and ventral sied in 1 case. The R. E.N.A.L score was 4.0-6.0, with an average of (5.0±0.8). The preoperative creatinine was 66.9-90.1μmol/L, with an average of (75.1±9.0)μmol/L, preoperative GFR of 44. 6- 67. 3 ml /min, with an average of(56.7±7.7)ml/min, preoperative hemoglobin level of 119.0-156.0g/L, with an average of (135.8±11.4)g/L. All patients underwent laparoscopic non-blocking partial nephrectomy assisted by 180w lateral green laser, free the surrounding area of the tumor fully and completely expose the renal tumor. The laser fiber was placed through the green laser hand piece, and the fiber was connected with normal saline to wash the strip. The initial green laser vaporization power was set at 80W, and the hemostasis power at 35W.About 3mm away from the edge of the tumor, and one optical fiber away from the renal parenchyma, the renal parenchyma was cut with 80W power. In order to reduce the interference by smoke, high-pressure flushing was used through the optical fiber while vaporizing, and an attractor was used to push and peel the tumor. In case of bleeding during operation, hemostatic power can be used to close the bleeding point and gradually advance until the tumor was completely removed. The wounds of renal inner medulla and renal outer cortex were continuously sutured in 1-3 layers with barbed suture. It involved 9 cases via retroperitoneal approach and 1 case via abdominal approach. The operation time, postoperative hemoglobin decrease, extraction time of negative pressure drainage, postoperative hospital stay, postoperative pathology and postoperative complications were recorded, and the serum creatinine level and GFR level of the affected side were followed up 1 month after operation.Results:All the operations were successfully completed, and there was no conversion to open surgery or radical nephrectomy. One case changed to scissors fast resection and sutured hemostasis due to severe intraoperative bleeding. The operation time was 90.0-120.0 min, with the average of (104.5±9.0)min. The postoperative hemoglobin level was 96.0-132.0g/L, with an average of (115.2±11.8)g/L, and the difference was statistically significant ( P<0.05). The postoperative hemoglobin decreased from 12.0g/L to 25.0g/L, with an average of (20.6±4.6)g/L. The time of vacuum drainage was 5.0-7.0 days, with an average of (5.7±0.7)d. Postoperative hospital stay was 6.0-8.0 days, with an average of (6.7±0.7)d. No bleeding, urinary leakage and other complications occurred in all patients. There were 7 cases of clear cell carcinoma, 2 cases of papillary renal cell carcinoma and 1 case of angiomyolipoma. All margins were negative. One month after operation, creatinine ranged from 66.0 to 90.4μmol/L, with an average of (76.8±8.3)μmol/L, which was not significantly different compared with that before operation ( P>0.05). One month after operation, GFR was 45.1-60.8 ml/min, and with an average of (55.5±4.7)ml/min, and there was no significant difference compared with preoperative data( P>0.05). Conclusions:For T 1aN 0M 0 stage and exophytic renal tumors, laparoscopic non-blocking partial nephrectomy assisted by lateral green laser is safe and effective.

Objective:To investigate clinical efficacy and safety of cultured autologous melanocyte transplantation for the treatment of non-segmental vitiligo accompanied by autoimmune thyroid diseases.Methods:From May 2008 to December 2018, a total of 2 284 patients with non-segmental vitiligo were retrospectively collected, who received cultured autologous melanocyte transplantation in Hangzhou Third People′s Hospital. Among these patients, 75 were also diagnosed with autoimmune thyroid diseases, including hyperthyroidism (42 cases) , hypothyroidism (18 cases) and Hashimoto′s thyroiditis (15 cases) . Efficacy and safety were compared between the vitiligo patients with autoimmune thyroid diseases (concomitant group) and those without (non-concomitant group) . Chi-square test was used to compare enumeration data.Results:Among the 2 284 patients, 1 085 were males and 1 199 were females, with an age of 25.0 ± 1.2 years and a disease duration of 5.1 ± 2.3 years. Six months after transplantation, 1 873 out of 2 209 patients in the non-concomitant group achieved favorable clinical response, with a response rate of 84.8%, including 1 162 achieving complete clinical response (52.6%) ; 46 out of 75 patients in the concomitant group achieved favorable clinical response, with a response rate of 61.3%, including 20 achieving complete clinical response (26.7%) ; the response rate and recovery rate were both significantly lower in the concomitant group than in the non-concomitant group ( χ2 = 29.72, 19.54, respectively, both P < 0.001) . Moreover, the response rate was significantly lower in the hypothyroidism group than in the hyperthyroidism group ( χ2 = 6.61, P = 0.010) . The incidence of isomorphic response at the donor site was significantly higher in the concomitant group than in the non-concomitant group (9.3% vs. 4.3%, χ2 = 4.31, P = 0.038) , so were the recurrence rates of vitiliginous patches at the recipient site after 1, 3, 5 and 10 years (concomitant group: 6.7%, 14.7%, 17.3%, 8.7%, respectively; non-concomitant group: 0.7%, 1.4%, 2.1%, 3.6%, respectively; χ2 = 29.96, 70.69, 67.23, 41.61, respectively, all P < 0.001) . Conclusion:Concomitant autoimmune thyroid diseases negatively affect the efficacy of cultured autologous melanocyte transplantation in the treatment of vitiligo, so effective measures should be taken to prevent isomorphic response and recurrence at the recipient site for patients with non-segmental vitiligo complicated by autoimmune thyroid diseases.