Objective:To construct a sizeable naive human Fab phage display antibody library to screen high-affinity specific antibodies in vitro. Methods:Total RNA was extracted from peripheral blood mononuclear cells (PBMCs) of 126 healthy individuals, subsequently reverse-transcribed into cDNA, and used as a template. PCR amplification was performed to obtain the V H from IgG, IgM and light chain κ, λ, separately, with the initial PCR products serving as templates for a second round of PCR. Overlap extension PCR was employed to generate fragments of the κ and λ light chains. These fragments were ligated with the phage vector pNC3, which harbors the variable region 1 of the heavy chain, to construct a recombinant phage plasmid. This plasmid was then electroporated into competent Escherichia Coli TG1 cells to establish a naive human Fab phage display antibody library. One hundred clones were randomly selected for identification and sequencing, and antibody gene polymorphisms were analyzed using the IMGT database and MAFFT software. Recombinant α-hemolysin from Staphylococcus aureus was utilized to screen Fab antibody fragments through biopanning of the antibody library, followed by random selection of phage ELISA-identified clones. The positive clones (antigen A450∶blank control A450≥2.1) were sequenced. Results:Two large naive Fab phage display antibody libraries were successfully constructed, in which the capacity of κ and λ chain antibody libraries were 1.25×10 11 and 1.54×10 11, respectively. The titers for two antibody libraries were 6.04×10 13 CFU/ml and 3.50×10 13 CFU/ml. The positive transformation insertion rates for κ and λ chain antibody libraries were 96% (96/100) and 100% (100/100), respectively. Sequence analysis revealed that all antibody sequences were unique. The amino acid sequences in the skeletal region were relatively conserved. In contrast, significant variations in the length of the complementarity determining region (CDR) were found, and the diversity of amino acid sequence of the complementary determining region was high, especially the CDR3. Analysis using the IMGT database indicated that the sequences exhibited a broad distribution across variable-diversity-joining gene families. After six rounds of panning, specific phage antibodies enrichment targeting α-hemolysin were achieved. A total of 142 monoclonal antibodies were sequenced, yielding 8 distinct Fab antibody sequences. Conclusion:This study successfully constructed two naive human Fab phage display antibody libraries with large capacity and good diversity, which can be used for screening human antibodies for serum epidemiology.

Objective:To construct a sizeable naive human Fab phage display antibody library to screen high-affinity specific antibodies in vitro. Methods:Total RNA was extracted from peripheral blood mononuclear cells (PBMCs) of 126 healthy individuals, subsequently reverse-transcribed into cDNA, and used as a template. PCR amplification was performed to obtain the V H from IgG, IgM and light chain κ, λ, separately, with the initial PCR products serving as templates for a second round of PCR. Overlap extension PCR was employed to generate fragments of the κ and λ light chains. These fragments were ligated with the phage vector pNC3, which harbors the variable region 1 of the heavy chain, to construct a recombinant phage plasmid. This plasmid was then electroporated into competent Escherichia Coli TG1 cells to establish a naive human Fab phage display antibody library. One hundred clones were randomly selected for identification and sequencing, and antibody gene polymorphisms were analyzed using the IMGT database and MAFFT software. Recombinant α-hemolysin from Staphylococcus aureus was utilized to screen Fab antibody fragments through biopanning of the antibody library, followed by random selection of phage ELISA-identified clones. The positive clones (antigen A450∶blank control A450≥2.1) were sequenced. Results:Two large naive Fab phage display antibody libraries were successfully constructed, in which the capacity of κ and λ chain antibody libraries were 1.25×10 11 and 1.54×10 11, respectively. The titers for two antibody libraries were 6.04×10 13 CFU/ml and 3.50×10 13 CFU/ml. The positive transformation insertion rates for κ and λ chain antibody libraries were 96% (96/100) and 100% (100/100), respectively. Sequence analysis revealed that all antibody sequences were unique. The amino acid sequences in the skeletal region were relatively conserved. In contrast, significant variations in the length of the complementarity determining region (CDR) were found, and the diversity of amino acid sequence of the complementary determining region was high, especially the CDR3. Analysis using the IMGT database indicated that the sequences exhibited a broad distribution across variable-diversity-joining gene families. After six rounds of panning, specific phage antibodies enrichment targeting α-hemolysin were achieved. A total of 142 monoclonal antibodies were sequenced, yielding 8 distinct Fab antibody sequences. Conclusion:This study successfully constructed two naive human Fab phage display antibody libraries with large capacity and good diversity, which can be used for screening human antibodies for serum epidemiology.

Objective:To explore the predictive value of miR-21 combined with high mobility group box-1 protein (HMGB1) for postoperative prognosis in patients with hypertensive intracerebral hemorrhage (HICH).Methods:A total of 137 HICH patients (HICH group) admitted to the Central Hospital of Yongzhou from March 2021 to March 2023 were retrospectively selected, and 60 healthy subjects undergoing physical examination in the same hospital during the same period were enrolled as the control group. Serum miR-21 and HMGB1 levels were compared between the two groups, and differences in serum miR-21 and HMGB1 levels among HICH patients with different disease severities and prognoses were analyzed. Univariate and multivariate logistic regression analyses were used to screen influencing factors for postoperative prognosis in HICH patients, and receiver operating characteristic (ROC) curve was applied to evaluate the predictive value of miR-21 combined with HMGB1 for postoperative prognosis.Results:The relative expression level of serum miR-21 on postoperative day 1 in the HICH group was significantly lower than that in the control group ( P<0.05), while the HMGB1 level was significantly higher ( P<0.05). With the aggravation of HICH severity, the relative expression of serum miR-21 significantly decreased (all P<0.05), while the HMGB1 level significantly increased (all P<0.05). The relative expression of serum miR-21 in the good prognosis group was significantly higher than that in the poor prognosis group ( P<0.05), while the HMGB1 level was significantly lower ( P<0.05). Univariate and multivariate logistic regression analyses showed that miR-21, HMGB1, preoperative Glasgow Coma Scale (GCS) score, and preoperative hematoma volume were influencing factors for postoperative prognosis in HICH patients (all P<0.05). ROC curve showed that the areas under the curve (AUC) of miR-21 and HMGB1 for predicting poor postoperative prognosis in HICH patients were 0.925 and 0.913, respectively, while the AUC of miR-21 combined with HMGB1 was 0.950. Conclusions:Peripheral blood miR-21 is significantly decreased and HMGB1 level is significantly increased in HICH patients. miR-21 combined with HMGB1 has a good predictive value for poor postoperative prognosis in patients with hypertensive intracerebral hemorrhage.

Objective:To explore the predictive value of miR-21 combined with high mobility group box-1 protein (HMGB1) for postoperative prognosis in patients with hypertensive intracerebral hemorrhage (HICH).Methods:A total of 137 HICH patients (HICH group) admitted to the Central Hospital of Yongzhou from March 2021 to March 2023 were retrospectively selected, and 60 healthy subjects undergoing physical examination in the same hospital during the same period were enrolled as the control group. Serum miR-21 and HMGB1 levels were compared between the two groups, and differences in serum miR-21 and HMGB1 levels among HICH patients with different disease severities and prognoses were analyzed. Univariate and multivariate logistic regression analyses were used to screen influencing factors for postoperative prognosis in HICH patients, and receiver operating characteristic (ROC) curve was applied to evaluate the predictive value of miR-21 combined with HMGB1 for postoperative prognosis.Results:The relative expression level of serum miR-21 on postoperative day 1 in the HICH group was significantly lower than that in the control group ( P<0.05), while the HMGB1 level was significantly higher ( P<0.05). With the aggravation of HICH severity, the relative expression of serum miR-21 significantly decreased (all P<0.05), while the HMGB1 level significantly increased (all P<0.05). The relative expression of serum miR-21 in the good prognosis group was significantly higher than that in the poor prognosis group ( P<0.05), while the HMGB1 level was significantly lower ( P<0.05). Univariate and multivariate logistic regression analyses showed that miR-21, HMGB1, preoperative Glasgow Coma Scale (GCS) score, and preoperative hematoma volume were influencing factors for postoperative prognosis in HICH patients (all P<0.05). ROC curve showed that the areas under the curve (AUC) of miR-21 and HMGB1 for predicting poor postoperative prognosis in HICH patients were 0.925 and 0.913, respectively, while the AUC of miR-21 combined with HMGB1 was 0.950. Conclusions:Peripheral blood miR-21 is significantly decreased and HMGB1 level is significantly increased in HICH patients. miR-21 combined with HMGB1 has a good predictive value for poor postoperative prognosis in patients with hypertensive intracerebral hemorrhage.

Objective:To investigate the incidence and prothrombotic risk factors of postoperative venous thromboembolism(VTE) in Cushing′s disease and to further develop an assessment model to identify those at high risk of postoperative VTE events.Methods:A retrospective study was performed in 82 patients who were admitted to Huashan Hospital, Fudan University during January 2019 and January 2020 and diagnosed with Cushing′s disease. These patients underwent the evaluation about their clinical, hormonal, and coagulation parameters, as well as ultrasonography and pulmonary angio-CT when necessary. The least absolute shrinkage and selection operator(LASSO) regression analysis was used to screen independent risk factors, and a nomogram model for postsurgical VTE risk assessment in Cushing′s disease was initially established, and Bootstrap method was used for internal verification. Finally, the predictive model was evaluated for calibration and clinical applicability in the study cohort.Results:Nineteen patients(23.17%) developed VTE events, with 14 cases occurring after endoscopic transsphenoidal surgery. Compared to patients without VTE, those in the VTE group were older( P<0.001), had longer postoperative bed rest, higher rates of current infection, higher HbA 1C levels, and more severe glucose tolerance impairment(all P<0.05). Through LASSO regression analysis, two independent risk factors for postoperative VTE were identified: Age and current infection. Then a VTE risk assessment nomogram model was established to predict the patients at high risk of VTE. In the nomogram model for VTE risk assessment, the area under the receiver operating characteristic curve was 0.868(95% CI 0.787-0.949), with the calibration curve closely aligning with the ideal diagonal line and the clinical decision curve exceeding the two extreme curves. Conclusions:Advanced perioperative assessment needs to be taken to screen those with high VTE risks in patients diagnosed with Cushing′s disease. Additionally, during the perioperative period, patients with Cushing′s disease should undergo mandatory physical activity or prophylactic anticoagulant therapy.

Traumatic cerebrospinal fluid leakage commonly presents in traumatic brain injury patients, and it may lead to complications such as meningitis, ventriculitis, brain abscess, subdural hematoma or tension pneumocephalus. When misdiagnosed or inappropriately treated, traumatic cerebrospinal fluid leakage may result in severe complications and may be life-threatening. Some traumatic cerebrospinal fluid leakage has concealed manifestations and is prone to misdiagnosis. Due to different sites and mechanisms of trauma and degree of cerebrospinal fluid leak, treatments for traumatic cerebrospinal fluid leakage varies greatly. Hence, the Craniocerebral Trauma Professional Group of Neurosurgery Branch of Chinese Medical Association and the Neurological Injury Professional Group of Trauma Branch of Chinese Medical Association organized relevant experts to formulate the " Chinese expert consensus on the diagnosis and treatment of traumatic cerebrospinal fluid leakage in adults ( version 2023)" based on existing clinical evidence and experience. The consensus consisted of 16 recommendations, covering the leakage diagnosis, localization, treatments, and intracranial infection prevention, so as to standardize the diagnosis and treatment of traumatic cerebrospinal fluid leakage and improve the overall prognosis of the patients.

Background::As a non-invasive and effective diagnostic method for small intestinal bacterial overgrowth (SIBO), wild-use of breath test (BT) has demonstrated a high comorbidity rate in patients with diarrhea-predominant irritable bowel syndrome (IBS-D) and SIBO. Patients overlapping with SIBO respond better to rifaximin therapy than those with IBS-D only. Gut microbiota plays a critical role in both of these two diseases. We aimed to determine the microbial difference between IBS-D overlapping with/without SIBO, and to study the underlying mechanism of its sensitivity to rifaximin.Methods::Patients with IBS-D were categorized as BT-negative (IBSN) and BT-positive (IBSP). Healthy volunteers (BT-negative) were enrolled as healthy control. The patients were clinically evaluated before and after rifaximin treatment (0.4 g bid, 4 weeks). Blood, intestine, and stool samples were collected for cytokine assessment and gut microbial analyses.Results::Clinical complaints and microbial abundance were significantly higher in IBSP than in IBSN. In contrast, severe systemic inflammation and more active bacterial invasion function that were associated with enrichment of opportunistic pathogens were seen in IBSN. The symptoms of IBSP patients were relieved in different degrees after therapy, but the symptoms of IBSN rarely changed. We also found that the presence of IBSN-enriched genera ( Enterobacter and Enterococcus) are unaffected by rifaximin therapy. Conclusions::IBS-D patients overlapping with SIBO showed noticeably different fecal microbial composition and function compared with IBS-D only. The better response to rifaximin in those comorbid patients might associate with their different gut microbiota, which suggests that BT is necessary before IBS-D diagnosis and use of rifaximin.Registration::Chinese Clinical Trial Registry, ChiCTR1800017911.

Objective:To understand the correlation between urinary iodine, salt iodine and thyroid nodules in children aged 8 - 10 years and pregnant women in different regions of Fujian Province, and to explore the influencing factors of thyroid diseases.Methods:A cross-sectional study was conducted in Dongshan County and Pingtan County of Fujian Province from September to November 2019. According to the inclusion criteria, 140 pregnant women and 270 children aged 8 - 10 years were selected in Dongshan County, and 189 pregnant women and 368 children aged 8 - 10 years were selected in Pingtan County. Random urine and edible salt samples were collected to determine iodine content, and iodine nutrition was evaluated in each population. Thyroid was examined by B ultrasound and questionnaire survey of thyroid diseases was carried out. The correlation between urinary iodine, salt iodine and thyroid nodules in children aged 8 - 10 years and pregnant women in different regions was analyzed.Results:There were statistically significant differences in the median urinary iodine among children in different genders (male: 151.30 μg/L, female: 130.30 μg/L) and regions (Dongshan County: 160.30 μg/L, Pingtan County: 129.70 μg/L, P < 0.05); there was no significant difference in the median urinary iodine among children of different ages (8, 9, 10 years old: 141.60, 128.05, 150.30 μg/L, P > 0.05). The median urinary iodine among pregnant women was 119.30 μg/L, and there was no significant difference in median urinary iodine among pregnant women in different stages and regions ( P > 0.05). The medians of salt iodine from children and pregnant women were 20.30 and 23.65 mg/kg, respectively. Urinary iodine in children was positively correlated with salt iodine ( r = 0.13, P < 0.05). However, there was no correlation between urinary iodine and salt iodine in pregnant women ( P > 0.05). The detection rate of thyroid nodules in children was 21.79% (139/638). There was significant difference in the detection rate of thyroid nodules in children of different ages ( P < 0.05). The detection rate of thyroid nodules in pregnant women was 4.26% (14/329). There was no correlation between detection rate of thyroid nodules and urinary iodine or salt iodine in children and pregnant women ( P > 0.05). Thyroid volume of children in the two counties was within the normal range, and there was no correlation between thyroid volume and urinary iodine or salt iodine ( P > 0.05). Conclusions:The iodine nutrition of children in Dongshan County and Pingtan County is suitable (100 - 199 μg/L), while iodine deficiency (< 150 μg/L) exists in pregnant women. Urinary iodine in children is related to salt iodine, and urinary iodine increased with increase of salt iodine. The prevalence of thyroid nodules in children of different ages is different, which requires further study. The detection rate of thyroid nodules in children and pregnant women is not correlated with urinary iodine and salt iodine.

Objective:To analyze the disease burden of iodine deficiency disorders (IDD) and its changes in China in 1990 and 2016.Methods:Based on the data of Global Burden of Disease Study 2016 (GBD 2016), a descriptive statistical method was used to compare and analyze the changes of deaths number, mortality, disability adjusted life year [DALY, including years of life lost (YLL) and years lived with disability (YLD)], DALY rate and other disease burden indicators caused by IDD in China in 1990 and 2016.Results:The GBD 2016 data showed that the deaths number from IDD in China decreased from 259.31 in 1990 to 116.41 in 2016, the mortality decreased from 0.022 827/100 000 in 1990 to 0.008 515/100 000 in 2016; and the mortality in different age groups had decreased, the children mortality in the < 5 years old group had decreased from 0.088 639/100 000 to 0.009 875/100 000. The DALY and YLD due to IDD in China increased from 406.13 thousand person-years and 391.68 thousand person-years in 1990 to 455.05 thousand person-years and 451.95 thousand person-years in 2016, while YLL decreased from 14.45 thousand person-years to 3.10 thousand person-years; the DALY rate, YLL rate, and YLD rate decreased from 35.75 person-years per 100 000, 1.27 person-years per 100 000 and 34.48 person-years per 100 000 in 1990 to 33.29 person-years per 100 000, 0.23 person-years per 100 000 and 33.06 person-years per 100 000 in 2016. The main component of DALY for IDD was YLD, and the proportion increased from 96.44% in 1990 to 99.32% in 2016.Conclusions:The mortality of IDD in China has decreased, and IDD in the younger age group has been effectively controlled; however, the disease burden, especially the burden caused by disability has increased, and the disability of the population due to IDD should be given special attention.

OBJECTIVE: To explore the effect of "diabetes specialists-community general practitioners-community nurse co-management mode" and "diabetes specialist management mode" on diabetic nephropathy (DN) in primary medical institutions. METHODS: Patients with type 2 diabetes admitted to Quanzijie Health Clinic of Jimusar County of Xinjiang Uygur Autonomous Region from October 2017 to March 2018 were enrolled. The Patients were divided into co-management group or specialist management group according to their administrative villages. The treatment plans of the two groups were formulated with reference to the current guidelines. The subjects of the co-management group were jointly managed by a fixed team composed of diabetes specialists from Jimusar Traditional Chinese Medicine Hospital, community general practitioners and community nurses from Quanzijie Health Clinic, and required to attend diabetes education courses every month. The diabetes specialist of Jimusar Traditional Chinese Medicine Hospital was responsible for the formulation and management of the treatment plan of the research object. Follow-up was fulfilled once every 4 weeks for 24 weeks in two groups. Before and after intervention, blood glucose, blood pressure, urinary albumin/creatinine ratio (UACR), estimated glomerular filtration rate (eGFR) as well as the utilization rate of angiotensin converting enzyme inhibitors/angiotensin II receptor blocker (ACEI/ARB) were collected. RESULTS: A total of 115 patients accomplished this study with 54 patients in co-management group and 61 patients in specialist management group. After 24 weeks of intervention, fasting glucose level, postprandial glucose level 2 hours after breakfast, glycosylated hemoglobin (HbA1c), Log UACR in co-management group and specialists management group were significantly decreased compared with baseline [fasting glucose level (mmol/L): 8.06±1.92 vs. 9.16±2.83, 8.21±2.10 vs. 9.06±1.89; postprandial glucose level 2 hours after breakfast (mmol/L): 12.26±3.78 vs. 14.11±5.28, 12.47±3.63 vs. 14.00±3.88; HbA1c: 0.074±0.014 vs. 0.082±0.023, 0.076±0.014 vs. 0.081±0.016; Log UACR (mg/g): 1.63±1.56 vs. 2.25±1.44, 1.84±1.65 vs. 2.43±1.56, all P < 0.05], but there was no statistical significance between the two groups [fasting glucose level (mmol/L): -1.10±0.47 vs. -0.85±0.36, postprandial glucose level 2 hours after breakfast (mmol/L): -1.85±0.88 vs. -1.53±0.68, HbA1c: -0.008±0.004 vs. -0.006±0.003, Log UACR (mg/g): -0.61±0.29 vs. -0.59±0.29, all P < 0.05]. There were no significant changes in blood pressure, serum creatinine and eGFR in the two groups before and after intervention. There were 18 and 24 patients with hypertension in co-management group and specialist management group, respectively. The utilization rates of ACEI/ARB in both groups after intervention were significantly higher than those before intervention [88.9% (16/18) vs. 22.2% (4/18), 95.8% (23/24) vs. 29.2% (7/24), both P < 0.01]. At the end of the study, the utilization rate of ACEI/ARB was similar between the two groups [88.9% (16/18) vs. 95.8% (23/24), P > 0.05]. CONCLUSIONS Both "diabetes specialists-community general practitioners-community nurse co-management mode" and "diabetes specialist management mode" can effectively decrease glucose levels and UACR levels of patients with type 2 diabetes as well as the standard use of antihypertensive agents, which has positive effects on the prevention and treatment on DN.
Blood Glucose ; Creatinine ; Diabetes Mellitus, Type 2 ; Diabetic Nephropathies ; Humans ; Prospective Studies