Objective: To develop a RHCE genotyping assay based on kompetitive allele-specific PCR (KASP) and assess its clinical accuracy for RhCE blood group determination. Methods: KASP primers were designed to interrogate three RHCE loci: the 109 bp insertion/deletion in intron 2, c. 307T>C, and c. 676C>G. A total of 1 194 RhD-positive inpatients from Chengdu were typed by both KASP genotyping and manual tube serology. Discordant samples (n=10) were retested by both methods and further resolved by Sanger sequencing. An additional 377 cases were tested for the c. 48C>G locus to evaluate the predictive accuracy of individual loci and combined locus testing for RhC antigen. Results: Genotyping concordance with serology was 100.0% for both the c. 676C>G locus (RhE/Rhe) and the c. 307T>C locus (Rhc). For RhC prediction using the 109 bp insertion, overall accuracy was 99.7% (1 191/1 194); the 3 discordant cases were confirmed by Sanger sequencing to be false negatives attributable to 109 bp deletion in intron 2. Testing the c. 48C>G allele for RhC prediction yielded 7 false positives, with an accuracy of 98.1% (370/377). RhC antigen status was determined by combining the 109 bp insertion and the c. 48C allele. After excluding 10 samples with inconsistent results between the two loci, the accuracy reached 100% in the remaining 367 samples. When both loci were applied in combination, accuracy reached 100% in the 367 cases with concordant results. Among the 1 194 patients, CCee (45.8%) and CcEe (31.7%) were the most common RhCE phenotypes. The e antigen had the highest positivity rate (92.2%), and the Ce haplotype was the most frequent (66.9%). Conclusion: The KASP-based RHCE genotyping method achieves high accuracy for clinical RhCE typing. Combining the 109 bp insertion/deletion with the c. 48C allele significantly improves RhC antigen prediction compared with either locus alone. This method was applied to RhCE genotyping of 1 194 RhD-positive inpatients in Chengdu, providing local RhCE phenotype and haplotype distribution data to support RhCE-matched transfusion practice.

Objective:To analyze the clinical characteristics of mannose phosphate isomerase-congenital disorders of glycosylation (MPI-CDG) and evaluated the outcomes following D-mannose treatment.Methods:This case-series study analyzed clinical manifestations, laboratory findings, imaging results, genetic data, and outcomes after D-mannose therapy in 5 children with MPI-CDG diagnosed at the Children′s Hospital of Fudan University between December 2014 and December 2024.Results:The age of onset ranged from 0.3 to 0.4 years in all 5 children, who initially presented with diarrhea and hypoglycemia. Associated manifestations included short stature (3 cases), anemia (3 cases), splenomegaly (3 cases), hepatomegaly (4 cases), elevated transaminases (4 cases), and hypoalbuminemia (4 cases). Liver pathology revealed hepatic fibrosis in 3 cases. Genetic testing identified 8 variants in the MPI gene, including 2 novel variants. Following D-mannose treatment, diarrhea and hypoglycemia resolved within 1-2 weeks in all children, with concurrent improvement in anemia. Notably except for Patient 1, who developed progressive splenomegaly, worsening hepatic fibrosis, and portal hypertension despite persistently normal transaminase and albumin levels, the other 4 children showed improvement in transaminase levels, resolution of hypoalbuminemia and amelioration of imaging abnormalities.Conclusions:MPI-CDG typically manifests in infancy with diarrhea and hypoglycemia, often accompanied by multi-system involvement. D-mannose treatment significantly improves metabolic abnormalities and most organ damages. However, close surveillance of liver status is warranted due to the risk of hepatic fibrosis progression in some cases.

Objective:To analyze the clinical characteristics of mannose phosphate isomerase-congenital disorders of glycosylation (MPI-CDG) and evaluated the outcomes following D-mannose treatment.Methods:This case-series study analyzed clinical manifestations, laboratory findings, imaging results, genetic data, and outcomes after D-mannose therapy in 5 children with MPI-CDG diagnosed at the Children′s Hospital of Fudan University between December 2014 and December 2024.Results:The age of onset ranged from 0.3 to 0.4 years in all 5 children, who initially presented with diarrhea and hypoglycemia. Associated manifestations included short stature (3 cases), anemia (3 cases), splenomegaly (3 cases), hepatomegaly (4 cases), elevated transaminases (4 cases), and hypoalbuminemia (4 cases). Liver pathology revealed hepatic fibrosis in 3 cases. Genetic testing identified 8 variants in the MPI gene, including 2 novel variants. Following D-mannose treatment, diarrhea and hypoglycemia resolved within 1-2 weeks in all children, with concurrent improvement in anemia. Notably except for Patient 1, who developed progressive splenomegaly, worsening hepatic fibrosis, and portal hypertension despite persistently normal transaminase and albumin levels, the other 4 children showed improvement in transaminase levels, resolution of hypoalbuminemia and amelioration of imaging abnormalities.Conclusions:MPI-CDG typically manifests in infancy with diarrhea and hypoglycemia, often accompanied by multi-system involvement. D-mannose treatment significantly improves metabolic abnormalities and most organ damages. However, close surveillance of liver status is warranted due to the risk of hepatic fibrosis progression in some cases.

Traumatic massive abdominopelvic hemorrhage presents significant clinical challenges due to its complex anatomy, deep-seated and predominantly non-compressible bleeding sites, difficulties in early recognition, and rapid disease progression. Patients are often complicated with shock and coagulopathy, making timely intervention extremely critical. Despite continuous advances in treatment techniques for trauma, clinical practice still faces significant challenges, including insensitive early warning systems, limited assessment means, suboptimal selection and timing of hemostatic interventions, insufficient multidisciplinary team (MDT) collaboration, and imperfect hemostasis strategy based on anatomical stratification. Focusing on the treatment chain of "early warning-assessment-hemostasis-support", the authors reviewed key challenges in managing massive abdominopelvic hemorrhage including the establishment of early warning criteria, standardization and intelligentization of focused assessment with sonography for trauma (FAST), standardized application of (REBOA), optimization of MDT collaborative mechanisms, and integration of anatomy-based hemostatic strategies, and proposed optimization pathways, aiming to improve the success rate in managing traumatic massive abdominopelvic hemorrhage.

Background With economic development and globalization, shift work has become prevalent across industries. Its relationship with type 2 diabetes mellitus (T2DM) attracts increasing attention. Objective To thoroughly explore the relationship between shift work and T2DM, and analyze the impacts of specific shift patterns on T2DM, so as to provide a basis for formulating reasonable shift schedules. Methods We conducted a 1:2 matched case-control study among adults (20-60 years) who ordered occupational health examinations at the Wuxi No.8 People's Hospital from November to December 2023. The case group comprised 200 T2DM patients, while the controls were 400 age-stratified matched non-diabetic individuals. General demographic characteristics, behavioral habits, medical history, and shift work exposure data (including shift patterns, frequency, and length of service) 5 years prior to diagnosis were collected through standardized questionnaires. Logistic regression adjusted for selected confounders was employed to evaluate the association between shift work and T2DM. Results The logistic regression analysis demonstrated that shift work was associated with an increased risk of T2DM. After adjusting for confounding factors, shift workers had a 3.55 times higher risk of being diagnosed T2DM compared to non-shift workers (OR=3.55, 95%CI: 1.026, 12.263). The risk varied across different shift patterns, and the three-shift two-rotation system showed the highest risk (OR=4.17, 95%CI: 1.921, 9.035), followed by the two-shift system (OR=2.94, 95%CI: 2.016, 4.281) and four-shift three-rotation system (OR=2.66, 95%CI: 1.611, 6.093). Workers with more than 3 monthly shift days had a 2.74-fold increased risk (95%CI: 1.658, 4.512) compared to non-shift workers. Additionally, working more than 8 h daily (OR=1.74, 95%CI: 1.185, 2.562) and having more than 20 years of service (OR=2.51, 95%CI: 1.581, 3.976) were both significantly associated with a higher T2DM risk. The trend tests revealed that each incremental increase in monthly shift days and length of service elevated T2DM risk by 2.61 times (95%CI: 1.813, 3.765) and 1.49 times (95%CI: 1.147, 1.931), respectively (P<0.05). Conclusion Shift work is an independent risk factor for T2DM, with three-shift two-rotation system posing the highest risk. Shift frequency, daily working hours, and length of service are all significant factors affecting the risk of T2DM. These findings support industry-specific shift policy reform and targeted glucose monitoring and health interventions are recommended for workers engaged in high-risk shift patterns (e.g., three-shift two-rotation system, frequent shifts) and those with prolonged shift work history (>20 years).

Traumatic massive abdominopelvic hemorrhage presents significant clinical challenges due to its complex anatomy, deep-seated and predominantly non-compressible bleeding sites, difficulties in early recognition, and rapid disease progression. Patients are often complicated with shock and coagulopathy, making timely intervention extremely critical. Despite continuous advances in treatment techniques for trauma, clinical practice still faces significant challenges, including insensitive early warning systems, limited assessment means, suboptimal selection and timing of hemostatic interventions, insufficient multidisciplinary team (MDT) collaboration, and imperfect hemostasis strategy based on anatomical stratification. Focusing on the treatment chain of "early warning-assessment-hemostasis-support", the authors reviewed key challenges in managing massive abdominopelvic hemorrhage including the establishment of early warning criteria, standardization and intelligentization of focused assessment with sonography for trauma (FAST), standardized application of (REBOA), optimization of MDT collaborative mechanisms, and integration of anatomy-based hemostatic strategies, and proposed optimization pathways, aiming to improve the success rate in managing traumatic massive abdominopelvic hemorrhage.

Glycosylation is one of the most important reactions in living organisms as it results in the formation of glycoconjugates with diverse biological functions. Sugar nucleotides are structurally composed of sugar and nucleoside diphosphate or monophosphate, which are widespread within a variety of biological cells. As glycosyl donors for the transglycosyl reactions catalyzed by Leloir-type glycosyltransferases, sugar nucleotides are essential for the synthesis of glycans and glycoconjugates. However, high costs and limited availability of nucleotide sugars prevent applications of biocatalytic cascades on an industrial scale. Therefore, attentions on synthetic strategies of sugar nucleotides have been increasing to achieve their wide applications in various fields. The 9 common sugar nucleotides in mammals have been fully studied with large-scale synthesis through chemical, enzymatic (chemo-enzymatic) and cell factory strategies. In addition to common sugar nucleotides, many rare sugar nucleotides are present in plants and bacteria. Although unnatural sugar nucleotides cannot be synthesized in organisms, they have great potential in research as substrates for glycosyltransferases in carbohydrate synthesis, as enzyme inhibitors in biochemical studies, and as components of glycoconjugate biosynthesis. Therefore, increasing attention has been paid to explore the efficient synthesis of unnatural sugar nucleotides. Currently, strategies for chemical synthesis of sugar nucleotides have been greatly improved, such as the use of effective catalysts for forming pyrophosphate bonds and the development of entirely new synthesis protocols. Multiple sugar nucleotides, especially unnatural sugar nucleotides, are synthesized chemically. However, chemical synthesis requires tedious protection and deprotection steps, resulting in complex steps, high cost and low yield. In contrast, enzymatic (chemo-enzymatic) and cell factory methods have significant advantages such as high yield, easy operation and easy process scale-up in the preparation of sugar nucleotides. Hence, they are prominent strategies for sugar nucleotide preparation. Herein, the biosynthesis and application of sugar nucleotides are reviewed, mainly focusing on the 9 sugar nucleotides common in mammals. The early strategies for enzymatic synthesis of sugar nucleotides generally used de novo synthesis pathway. With the discoveries of enzymes involved in salvage pathway of sugar nucleotide synthesis and the development of one-pot multienzyme (OPME) method, the synthesis of sugar nucleotides was greatly simplified. Cell factory method employs the microbial living cells as a “processing plant” by engineering their metabolic pathways through genetic engineering technology. The cell factory method has high yield, and has been applied for efficient synthesis of several sugar nucleotides. Moreover, the strategy of gram-scale synthesis of multiple rare sugar nucleotides by cascade reactions from common sugar nucleotides using sugar nucleotides synthases cloned from different sources was illustrated. In recent years, the synthesis cost of sugar nucleotides has been further reduced through various ways, such as regeneration of nucleotides, regeneration of organic cofactors, and application of immobilized enzyme technology. Furthermore, through the continuous improvement of sugar nucleotide purification process, the use of high concentration of multi-enzyme cascade and rapid non-chromatographic purification process, the synthesis of multiple sugar nucleotides and their derivatives from monosaccharides was achieved, which gradually broke the limitations of the existing strategy. With the efficient synthesis of sugar nucleotides, their applications in various fields have been increasingly explored, including the synthesis of glycans and glycoconjugates, biochemical characterization of glycosyltransferases and bioorthogonal labeling strategies, which are of great significance to the research of biochemistry, glycobiology and the development of related pharmaceutical products.

Objective:To study the mechanism of electroacupuncture (EA) on improving insulin resistance and in vitro fertilization and embryo transfer (IVF-ET) pregnancy outcome in polycystic ovary syndrome (PCOS) patients with phlegm-damp syndrome based on DNA methylation and insulin metabolism signaling pathway. Methods:In this prospective randomized controlled study, 100 PCOS patients with phlegm-damp syndrome who underwent IVF-ET in Department of Reproduction and Genetics in Affiliated Hospital of Shandong University of Traditional Chinese Medicine from January 2020 to December 2022 were enrolled and divided into a treatment group (EA therapy) and control group (placebo needling) by random number table, 50 patients in each group. Fixed antagonist regimen was used to promote ovulation in the two groups. Patients received EA therapy and placebo needling respectively twice a week from the menstrual cycle before oocyte retrieval till human chorionic gonadotrophin injection day. The granulosa cells were collected. The improvement of phlegm and dampness syndrome, homeostasis model assessment insulin resistance (HOMA-IR), dosage and number of days of gonadotropins (Gn) used, number of oocytes retrieved and high-quality embryos, fertilization rate, clinical pregnancy rate, early miscarriage rate and live birth rate of patients in the two groups were compared. Bisulfite amplicon sequencing was used to evaluate the methylation levels of the INSR gene promoter region in ovarian granulosa cells of patients in the two groups. Real-time polymerase chain reaction and Western blotting technology were used to detect the expression of INSR, PI3K and GLUT4. Results:The phlegm-dampness score (15.23±1.57) and HOMA-IR (2.82±0.39) of the experimental group after treatment were significantly lower than those before treatment (21.65±3.61 and 3.34±0.56), and the differences were statistically significant ( P<0.001, P=0.014). The differences of the phlegm-dampness score (-5.76±2.86) and HOMA-IR (-2.67±0.06) before and after treatment in the experimental group were significantly greater than those in control group (-1.64±0.84, -0.11±0.04), and the differences were all statistically significant ( P<0.001, P=0.021). In the experimental group, the dosage of Gn used by patients [(2 119.53±338.28) U] and the duration of Gn used [(10.16±1.25) d] were significantly reduced compared with control group [(2 405.65±434.20) U, P=0.005; (10.94±1.46) d, P=0.026], and the number of high-quality embryos (3.54±1.04) was significantly increased compared with control group (2.66±1.87, P=0.014). Fertilization rate [66.91% (552/825)], clinical pregnancy rate [63.27% (31/49)] and live birth rate [51.02% (25/49)] were significantly higher in the experimental group than in control group [60.20% (475/789), 41.67% (20/48), 31.25% (15/48)], and the differences were all statistically significant ( P=0.005, P=0.033, P=0.048). There were no significant differences in the number of oocytes retrieved and early abortion rate between the two groups (all P>0.05). The methylation levels of INSR gene promoter sites 38, 47, 56, 59, 94 and 143 in the experimental group were significantly lower than those in control group (all P<0.05), but there were no significant differences in methylation levels of sites 71, 74, 154, 156 and 162 between the two groups (all P>0.05). Compared with control group, the mRNA and protein expression of INSR, PI3K and GLUT4 was up-regulated in the experimental group (all P<0.05). Conclusion:EA may improve IR by down-regulating INSR methylation levels and up-regulating INSR, PI3K and GLUT4 expression, thereby enhancing the quality of embryos and improving the pregnancy outcome in PCOS patients.

Objective:To study the mechanism of electroacupuncture (EA) on improving insulin resistance and in vitro fertilization and embryo transfer (IVF-ET) pregnancy outcome in polycystic ovary syndrome (PCOS) patients with phlegm-damp syndrome based on DNA methylation and insulin metabolism signaling pathway. Methods:In this prospective randomized controlled study, 100 PCOS patients with phlegm-damp syndrome who underwent IVF-ET in Department of Reproduction and Genetics in Affiliated Hospital of Shandong University of Traditional Chinese Medicine from January 2020 to December 2022 were enrolled and divided into a treatment group (EA therapy) and control group (placebo needling) by random number table, 50 patients in each group. Fixed antagonist regimen was used to promote ovulation in the two groups. Patients received EA therapy and placebo needling respectively twice a week from the menstrual cycle before oocyte retrieval till human chorionic gonadotrophin injection day. The granulosa cells were collected. The improvement of phlegm and dampness syndrome, homeostasis model assessment insulin resistance (HOMA-IR), dosage and number of days of gonadotropins (Gn) used, number of oocytes retrieved and high-quality embryos, fertilization rate, clinical pregnancy rate, early miscarriage rate and live birth rate of patients in the two groups were compared. Bisulfite amplicon sequencing was used to evaluate the methylation levels of the INSR gene promoter region in ovarian granulosa cells of patients in the two groups. Real-time polymerase chain reaction and Western blotting technology were used to detect the expression of INSR, PI3K and GLUT4. Results:The phlegm-dampness score (15.23±1.57) and HOMA-IR (2.82±0.39) of the experimental group after treatment were significantly lower than those before treatment (21.65±3.61 and 3.34±0.56), and the differences were statistically significant ( P<0.001, P=0.014). The differences of the phlegm-dampness score (-5.76±2.86) and HOMA-IR (-2.67±0.06) before and after treatment in the experimental group were significantly greater than those in control group (-1.64±0.84, -0.11±0.04), and the differences were all statistically significant ( P<0.001, P=0.021). In the experimental group, the dosage of Gn used by patients [(2 119.53±338.28) U] and the duration of Gn used [(10.16±1.25) d] were significantly reduced compared with control group [(2 405.65±434.20) U, P=0.005; (10.94±1.46) d, P=0.026], and the number of high-quality embryos (3.54±1.04) was significantly increased compared with control group (2.66±1.87, P=0.014). Fertilization rate [66.91% (552/825)], clinical pregnancy rate [63.27% (31/49)] and live birth rate [51.02% (25/49)] were significantly higher in the experimental group than in control group [60.20% (475/789), 41.67% (20/48), 31.25% (15/48)], and the differences were all statistically significant ( P=0.005, P=0.033, P=0.048). There were no significant differences in the number of oocytes retrieved and early abortion rate between the two groups (all P>0.05). The methylation levels of INSR gene promoter sites 38, 47, 56, 59, 94 and 143 in the experimental group were significantly lower than those in control group (all P<0.05), but there were no significant differences in methylation levels of sites 71, 74, 154, 156 and 162 between the two groups (all P>0.05). Compared with control group, the mRNA and protein expression of INSR, PI3K and GLUT4 was up-regulated in the experimental group (all P<0.05). Conclusion:EA may improve IR by down-regulating INSR methylation levels and up-regulating INSR, PI3K and GLUT4 expression, thereby enhancing the quality of embryos and improving the pregnancy outcome in PCOS patients.

Female ; Humans ; Anti-Mullerian Hormone ; Sexual Development ; Peptide Hormones