To enhance the scientific research participation of the medical postgraduates, and pro-mote core competitiveness of the medical colleges, based on analysis of the necessity of patent application in medical colleges postgraduate, we constructed the teaching teams including the medical professors, patent engineers, graduate teaching manager, and science and technology managers. The problem-based learning (PBL) and case-based learning (CBL) as teaching methods were used in practice. The patent courses included the reference search and analysis, basic knowledge of patent law, and patent application training module were constructed; and the teaching effect were evaluated and optimized through the scores of the patent basic theory test, research output, and the training of the patent application. The patent course of the medi-cal postgraduates provides a reference for cultivating the compound talents have scientific research innova-tion and patent application capability.

Abstract: Objective　The aim of this study was to observe the expression levels and clinical significance of peripheral blood interferon γ-inducible protein-10 (IP-10) and various cytokines in patients with different infection statuses of tuberculosis and to assess the efficacy of latent tuberculosis infection (LTBI) in the progression to active tuberculosis (ATB). Methods　Seventy-six outpatient and inpatient cases from the Third People's Hospital of Kunming were collected and analyzed from March 2023 to February 2024. The patients were divided into three groups: ATB group (31 cases, 17 males, median age 33 years), LTBI group (27 cases, 17 males, median age 29 years), and healthy control (HC) group (18 cases, 11 males, median age 25 years). Peripheral blood samples from the three groups were taken and the expression levels of IP-10 and cytokines IL-6, IL-4, IL-8, IL-10, IL-12, IL-2, and TNF-α were detected using enzyme-linked immunosorbent assay (ELISA) methods. The t-test was used for normally distributed samples, while the Mann-Whitney U test was used for skewed distributions. For comparisons between multiple groups, the Kruskal-Wallis H test was first employed, followed by Dunn's multiple comparison test for pairwise comparisons. Finally, the effectiveness of each cytokine in distinguishing different population groups was analyzed. Results　The expression levels of peripheral blood IP-10 were higher in the LTBI and ATB groups than in the HC group, but the area under the curve (AUC) of the receiver operating characteristic (ROC) of the subjects showed moderate sensitivity (AUC:0.7-0.9) and low specificity (AUC:0.5-0.7). The IL-6 expression levels were in the order of high to low in the ATB group, LTBI group, and HC group, where the HC group was significantly lower than the ATB and LTBI groups (F=12.15, P<0.001). The sensitivity and specificity of the ATB group were higher than those in the HC group. Conclusions　IP-10 exhibits unique advantages in distinguishing different tuberculosis statuses. The predictive efficacy of a single cytokine is limited. Combining multiple cytokines such as IL-6 with clinical manifestations, a more accurate and comprehensive prediction model can be established.

Objective To study the sensitivity and specificity of a double monoclonal antibody sandwich enzyme-linked immunosorbent assay (DMcAbS-ELISA)for the detection of F1 antigen of Yersinia pestis (Y.pestis).Methods Viscera (viz.liver and spleen)specimens of infected mice with virulent Y.pestis and negative control mice were detected by bacteriological test,DMcAbS-ELISA and reverse indirect hemagglutination assay (RIHA) for the F1 antigen.Results The 225 control specimens were all negative tested by plague bacteriology testing,DMcAbS-ELISA and RIHA.A total of 308 plague-infected mouse organ specimens were tested,and the positive detection rate was 92.21％ (284/308),90.91％(280/308) and 89.61％ (276/308),respectively,with germiculture,DMcAbS-ELISA and RIHA,and the difference was not statistically significant(x2=5.65,P＞0.05).The coincidence rate of DMcAbS-ELISA and bacterial culture was 97.00％[(274+243)/533],Kappa =0.940;RIHA in line with the rate was 99.25％[(276+253)/533],Kappa =0.985.Authenticity comparison of F1 antigen detection in viscera specimens:sensitivity,specificity,positive predictive value,negative predictive value,adjusted agreement and Youden's index was 96.48％(274/284),97.59％(243/249),97.86％ (274/280),96.05 ％(243/253),96.99％[1/4×(274/280+274/284+243/253+243/249)]and 0.9407,respectively,for DMcAbS-ELISA and 96.13％(273/284),98.80％(246/249),98.91％(273/276),95.72％(246/257),97.39％[1/4×(273/276+273/284+246/257±246/249)]and 0.9492,respectively,for RIHA.The detection sensitivity of DMcAbS-ELISA and RIHA was 2.7×104 cfu/ml and 2.2×105 cfu/ml,for Y.pestis,respectively,and was 10 μg/L for F1 antigen.Conclusions DMcAbS-ELISA assay is a sensitive,specific,simple and fast method for detection of the F1 antigen,and it has a potential application value in rapid diagnosis of plague.

OBJECTIVETo investigate lyophilization of SM-SLN.

METHODThe parameters of lyophilization process was optimized. In addition, the protective effect of various types and concentrations of cryoprotectants were tested by shape, colour and disparity.

RESULTThe mixture of 2% lactose and 2% glucose could better prevent nanoparticles from aggregating, the optimal lyophilization process was followed: precooled at -45 degrees C for 10 hr; primary drying at -25 degrees C for 5 hr; secondary drying at 10 degrees C for 3 hr; finally drying at 30 degrees C for 6 hr.

CONCLUSIONChanges in particle size distribution during lyophilization could be minimized by optimizing the parameters of the lyophilization process and adding supporting agent.

Drug Carriers ; chemistry ; Freeze Drying ; methods ; Glucose ; chemistry ; Lactose ; chemistry ; Lipids ; chemistry ; Milk Thistle ; chemistry ; Nanotechnology ; Particle Size ; Plants, Medicinal ; chemistry ; Silymarin ; administration & dosage ; chemistry ; isolation & purification ; Technology, Pharmaceutical ; methods

Objective To assess the quantitative relationship between the distribution of Himalayan marmot and its ecological environment,the terrain,the temperature and the precipitation,using remote sensing and geographic information system in Qinghai province.Methods The distribution of Himalayan marmot was located by Google Earth and ArcGIS software and by using field survey data provided by Chinese Center for Disease Control and Prevention.The corresponding ecological environment of marmot including terrain,temperature and precipitation were derived from the spatial information datasets.All results were processed according to the overlay and statistics analysis using ArcGIS software.Results Seventy-seven point twenty-seven percent(153/198) of Himalayan marmot were distributed in the area of elevation between 3000 and 4000 meters.The number of marmot reached the highest when the slope was between 0 and 17 degrees,and aspect range was between 91 and 270 degrees,180 degree was as south direction.During the period with the maximum temperature of the warmest month of 14.3-17.5 ℃,17.6-20.8 ℃ and 20.9-24.0 ℃,the distribution of marmot reached 95％(186/198) of the total area.Meanwhile,most of the marmot were presented in the area with average precipitation of 46-108 mm.Conclusions A quantitative analysis of appropriate ecological environment of Himalayan marmot in a large scope is carried uul successfully using remote sensing and geographic information system.The study indicates that spatial information technology has important applications in plague prevention and control.

Abstract: To explore the clinical characteristics, diagnosis and treatment of imported severe malaria and COVID-19 co-infection cases, and to provide scientific basis for epidemic prevention and control measures. The epidemiological characteristics, clinical manifestations, laboratory tests, treatment process and prognosis of 4 cases of severe malaria and COVID-19 co-infection with confirmed diagnosis were analyzed retrospectively. Four cases of severe malaria were African returnees of the same batch, male, aged 40-54 years old, with the same journey track. They all had African work and life history and acute onset. The main clinical manifestations were fever (4/4), chills (3/4), chills (3/4), nausea and vomiting (3/4), diarrhea (4/4), fatigue and anorexia (4/4). Two cases had headache and dizziness, confusion, muscle aches, two cases had cough, one cases had sputum, sore throat and runny urine. All 4 cases were confirmed by positive nucleic acid detection of the new coronavirus (2019-nCOV) in throat swabs. Plasmodium falciparum was found by microscopic examination of peripheral blood smears of all patients, and all of them were consistent with high altitude helminthiasis. All cases were accompanied by abnormal liver function and severe hypoproteinemia, two cases were hyperbilirubinemia, three cases were dyslipidemia, three cases were involved in abnormal tertiary hemogram with different degrees of elevation of procalcitonin, two cases were lactic acid poisoning, and one case was hypoglycemia. One case showed viral pneumonia on chest CT. All cases were treated individually according to the different conditions and were discharged after improvement, and were rechecked for 2019-nCOV nucleic acid and microscopic examination of blood smear negative for Plasmodium.During the global COVID-19 epidemic, the emergence of coinfection cases of con-infection of imported malaria parasites and severe acuterespiratory syndrome coronavirus 2 (SARS-CoV-2) makes the clinical diagnosis and treatment more complicated. It is important to establish the awareness of simultaneous prevention and diagnosis of COVID-19 and malaria for local prevention and control and early warning of severe cases, and timely and effective formulation of treatment plan to improve the comprehensive treatment efficiency.

Objective To further study the regulatory mechanisms of uncarboxylated osteocalcin on the glucose metabolism,rat uncarboxylated osteocalcin fusion protein was expressed,purified and identified.Methods The rat osteocalcin gene (BGLAP) was cloned into pSumo-Mut expression vector.The pSumo-BGLAP plasmid was obtained and identified.Through transformed in prokaryotic host bacterium and induced by isopropyl 3-D-thiogalactoside (IPTG) and 11 ℃ low temperature,the Sumo-BGLAP fusion protein was expressed.The purification and activity identification of the fusion protein was performed by Ni2+ affinity chromatography and insulin secretion experiments,respectively.Results The enzyme digestion and sequencing results of the plasmid showed that the recombinant plasmid pSumo-BGLAP was constructed successfully,which matched the target gene and protein.After the transformation,induction and purification,the high purity Sumo-BGLAP fusion protein was obtained.The data from insulin secretion experiments show that the fusion protein can promote insulin secretion under 16.7 mmol · L-1 glucose conditions [(37.64 ± 3.80)μU· L-1at 16.7 mmol · L-1glucose,(63.91 ±4.67) μU · L-1 at 16.7 mmol · L-1 glucose +0.03 ng· mL-1 Sumo-BGLAP,(68.47 ±5.83) μU · L-1 at 16.7 mmol · L-1 glucose +0.3 ng· mL-1 Sumo-BGLAP,P ＜ 0.01)].Conclusion The rat uncarboxylated osteocalcin fusion protein is successfully achieved and purified.The fusion protein with biological activity can be used to further study the regulatory mechanisms of uncarboxylated osteocalcin on the glucose metabolism.

AIM To establish a UFLC-QTRAP-MS/MS method for the simultaneous content determination of phenylalanine,isoleucine,methionine,valine,proline,tyrosine,aspartic acid,histidine,arginine,lysine and glutamic acid in Gynostemma pentaphyllum (Thunb.) Makino.METHODS The analysis of aqueous extract of G.pentaphyllum was performed on a Waters XBridge Amide column (2.1 mm × 100 mm,3.5 μm),with the mobile phase comprising of water-acetonitrile (containing 0.2％ formic acid) flowing at 0.6 mL/min in a gradient elution manner.RESULTS Eleven amino acids showed good linear relationships within their own ranges (r ＞0.998 9),whose average recoveries were 90.74％-103.05％ with the RSDs of 0.61％-5.00％.CONCLU-SION This accurate,stable and reproducible method can be used for the quality control of G.pentaphyllum.

Objective To construct a human tumor necrosis factor (hTNF-a) plasmid and identify it to optimize the fermentation conditions of hTNF-α protein so as to achieve high expression in Escherichia coli.Methods The gene of hTNF-a was cloned into pET24a vector to obtain the pET24a-hTNF-a expression plasmid that was transformed into Escherichia coli BL21(DE3),and the expression conditions of BL21 (DE3) were optimized.Results The plasmid of pET24a-hTNF-α was successfully constructed and identified by PCR and digestion,which was consistent with the target fragment hTNF-α.The plasmid was transformed into Escherichia coli BL21(DE3),the best induced expression conditions of Escherichia coli BL21 (DE3) were as follows:M9+LB medium,37℃,0.5 mmol/L IPTG,pH =7.5,and induction time was 5 h.The results showed that dry weight of the cells and the rate of TNF were increased by 2.56 times and 3.68 times,respectively,and the expression rate of hTNF-α was increased by 3.49 times from 9.38％ to 32.74％.Conclusion The optimal conditions for the expression of plasmid pET24a-hTNF-α in Escherichia coli were determined.

Objective To investigate the frequency of CD4~+CD25~+FoxP3~+regulatory T cells (Treg)and the expression of the functional protein,FoxP3,in patients with active tuberculosis and the relationship between Treg and the pathogenesis of tuberculosis.Methods Forty-five patients with active tuberculosis(including 25 cases of pulmonary tuberculosis and 20 tuberculous lymphadenitis), 20 healthy controls,20 recovered tuberculosis patients and 6 patients with reactive hyperplasia in cer- vical lymph node were enrolled.The frequency of CD4~+ CD25~+ FoxP3~+ Treg in the peripheral blood was measured by flow cytometry.FoxP3 mRNA expression was determined by real-time reverse transcriptase-polymerase chain reaction(RT-PCR)and the expression of FoxP3 protein in lymphoid tissues was measured by immunohistochemistry.Results The frequency of natural Treg in the peripheral blood from the patients with active tuberculosis was 2.91%?0.23%,which was signifi- cantly higher than that of healthy control group(1.22%?0.18%)and recovered tuberculosis patients(1.50%?0.17%,P