The mutations in human disease-causing genes are predominantly caused by point mutations, with more than half of them being transitions between guanine (G) and adenine (A). Precise and efficient in situ repair of these mutations is the most ideal approach for the treatment of genetic diseases. Given that most point mutations are transitions between G and A, adenine base editors (ABEs) based on the CRISPR/Cas9 system, which convert A to G, are particularly important for repairing these mutations in the treatment of human genetic diseases. In recent years, ABEs have been continuously optimized, with both activity and fidelity being improved. Here we summarize the progress of ABEs, especially those key mutants developed during the process of ABE optimization. It also reflects on the existing defects in current ABEs. Additionally, the article reviews the clinical applications (including preclinical studies) of ABE. Overall, the article aims to provide references for the discovery and optimization of new ABEs and their applications.

Objective:To explore the early effects and safety of using a hybrid fixation strategy with 3D-printed porous tantalum metal augments to reconstruct substantial bone defects in complex primary total knee arthroplasty (TKA).Methods:A retrospective analysis was performed on the clinical data from August 2019 to September 2023, encompassing 20 patients (21 knees) with significant bone loss who underwent hybrid fixation with 3D-printed porous tantalum augments. The procedures were conducted at two medical centers: the First Affiliated Hospital, School of Medicine, Zhejiang University (11 cases) and Xiangya Hospital of Central South University (9 cases). The study cohort comprised 6 males (6 knees) and 14 females (15 knees), with a mean age of 61.05±11.23 years (range, 42-80 years). The distribution of cases was 7 on the left side and 14 on the right side. All cases were categorized as type 3 according to the Anderson Orthopaedic Research Institute (AORI) classification system. The cohort included 19 unilateral and 1 bilateral case, with 5 involving complex primary replacements (3 with Charcot arthropathy, 1 with syphilitic arthropathy, and 1 with severe valgus deformity) and 16 revision surgeries (13 for aseptic loosening and 3 for infection). Preoperative assessments included routine CT scans and digital three-dimensional reconstructions to identify large metaphyseal defects exceeding 50% of the metaphyseal area or those thicker than 10 mm. For such defects, 3D-printed standardized porous tantalum augments were implemented. In cases of extensive cavitary bone defects or severe metaphyseal defects where the medial and lateral defects collectively exceeded 80% of the metaphyseal region or where the residual bone stock was insufficient for screw fixation of standardized augments, 3D-printed personalized custom-made porous tantalum augments were employed for hybrid fixation and repair. Comparative analyses were conducted on pre- and postoperative imaging data (prosthesis positioning and complications), knee range of motion (ROM), visual analogue scale (VAS) for pain, and Knee Society score (KSS).Results:Of the cases, 17 were repaired using standardized 3D-printed porous tantalum augments, while 4 underwent repairs with customized augments for hybrid fixation. Follow-up averaged 26.5±15.0 months (range, 12-62 months). There was a significant increase in knee ROM, improving from 72.8°±31.9° preoperatively to 113.2°±6.8° at 12 months postoperatively ( P<0.05). VAS scores decreased from 6.6±1.4 preoperatively to 2.5±1.0 at 12 months postoperatively ( P<0.05). Similarly, KSS improved from 52.8±6.4 preoperatively to 80.7±7.9 at 12 months postoperatively ( P<0.05). There were no incidences of prosthesis displacement, poor bone integration, or postoperative infections. Conclusion:The hybrid fixation strategy employing 3D-printed porous tantalum augments has been found to be effective in addressing significant bone defects in TKA. The follow-up results indicate a satisfactory biological integration of the porous tantalum metal augments with the host bone, which has resulted in substantial improvements in pain relief and knee joint functionality.

Objective:To investigate the influencing factors of participation in free health check-up among community residents.Methods:From 2012 to 2022 Xinzhuang Community Health Service Center of Shanghai Minhang district provided 5 free health check-up for local residents, once every 2 years. Among 5 904 eligible community residents with a mean age of (66.01±5.87) years, 682 (11.55 %), 912 (15.45 %), 842 (14.26 %), 934 (15.82 %), 1 061 (17.97 %) and 1 473 (24.95 %) participated in 5, 4, 3, 2, 1 and 0 health check-ups during 10 years, respectively. The influencing factors of participant frequency were analyzed with multivariate logistic regression model.Results:Multivariate logistic regression analysis showed that age 65 years and above ( OR=0.685, 95% CI: 0.625-0.751, P<0.001), exercising once a week or more ( OR=1.142, 95% CI: 1.031-1.266, P=0.011), and underweight ( OR=0.665, 95% CI:0.496-0.891, P=0.006) were independent factors influencing the participant in free health check-up among community residents. Conclusion:Community residents with older age or underweight are less likely to participate free health check-up, while those with frequent exercise like to participate.

Objective To investigate the incidence rate,clinical characteristics,and prognosis of neonatal stroke in Shenzhen,China.Methods Led by Shenzhen Children's Hospital,the Shenzhen Neonatal Data Collaboration Network organized 21 institutions to collect 36 cases of neonatal stroke from January 2020 to December 2022.The incidence,clinical characteristics,treatment,and prognosis of neonatal stroke in Shenzhen were analyzed.Results The incidence rate of neonatal stroke in 21 hospitals from 2020 to 2022 was 1/15 137,1/6 060,and 1/7 704,respectively.Ischemic stroke accounted for 75％(27/36);boys accounted for 64％(23/36).Among the 36 neonates,31(86％)had disease onset within 3 days after birth,and 19(53％)had convulsion as the initial presentation.Cerebral MRI showed that 22 neonates(61％)had left cerebral infarction and 13(36％)had basal ganglia infarction.Magnetic resonance angiography was performed for 12 neonates,among whom 9(75％)had involvement of the middle cerebral artery.Electroencephalography was performed for 29 neonates,with sharp waves in 21 neonates(72％)and seizures in 10 neonates(34％).Symptomatic/supportive treatment varied across different hospitals.Neonatal Behavioral Neurological Assessment was performed for 12 neonates(33％,12/36),with a mean score of(32±4)points.The prognosis of 27 neonates was followed up to around 12 months of age,with 44％(12/27)of the neonates having a good prognosis.Conclusions Ischemic stroke is the main type of neonatal stroke,often with convulsions as the initial presentation,involvement of the middle cerebral artery,sharp waves on electroencephalography,and a relatively low neurodevelopment score.Symptomatic/supportive treatment is the main treatment method,and some neonates tend to have a poor prognosis.

Objective To investigate whether activation of mitochondrial acetal dehydrogenase 2(ALDH2)alleviates hypoxic pulmonary hypertension by regulating the SIRT1/PGC-1α signaling pathway.Methods Thirty 8-week-old C57 BL/6 mice were randomized into control,hypoxia,and hypoxia+Alda-1(an ALDH2 activator)group(n=10),and the mice in the latter two groups,along with 10 ALDH2 knockout(ALDH2-/-)mice,were exposed to hypoxia(10%O2,90%N2)with or without daily intraperitoneal injection of Alda-1 for 4 weeks.The changes in right ventricular function and pressure(RVSP)of the mice were evaluated by echocardiography and right ventricular catheter test,and pulmonary artery pressure was estimated based on RVSP.Pulmonary vascular remodeling,right ventricular injury,myocardial α-SMA expression,distal pulmonary arteriole muscle normalization,right ventricular cross-sectional area,myocardial cell hypertrophy,and right cardiac hypertrophy index were assessed with HE staining,immunofluorescence staining and WGA staining,and the expressions of ALDH2,SIRT1,PGC-1α,P16INK4A and P21CIP1 were detected.In pulmonary artery smooth muscle cells with hypoxic exposure,the effect of Alda-1 and EX527 on cell senescence and protein expressions was evaluated using β-galactose staining and Western blotting.Results The wild-type mice with hypoxic exposure showed significantly increased RVSP,right ventricular free wall thickness and myocardial expressions of P16INK4A and P21CIP1,which were effectively lowered by treatment with Alda-1 but further increased in ALDH2-/-mice.In cultured pulmonary artery smooth muscle cells,hypoxic exposure significantly increased senescent cell percentage and cellular expressions of P16INK4A and P21CIP1,which were all lowered by treatment with Alda-1,but its effect was obviously attenuated by EX527 treatment.Conclusion ALDH2 alleviates hypoxia-induced senescence of pulmonary artery smooth muscle cells by upregulating the SIRT1/PGC-1α signaling pathway to alleviate pulmonary hypertension in mice.

Objective:To discuss the effect of D-limonene on the proliferation and apoptosis of the glioblastoma(GBM)cells,and to clarify its possible mechanism.Methods:The GBM cells were divided into control group(0 mmol·L-1 D-limonene)and 0.2,0.4,0.6,0.8,and 1.0 mmol·L-1 D-limonene groups.CCK-8 method was used to detect the inhibitory rates of proliferation of the cells in various groups;clone formation assay was used to detect the clone formation rates of the cells in various groups;Annexin Ⅴ-FITC/PI method was used to detect the apoptotic rates of the cells in various groups;Western blotting method was used to detect the expression levels of protein kinase B(AKT),B-cell lymphoma-2(Bcl-2),Bcl-2-associated X protein(Bax),and poly adenosine diphosphate(ADP)-ribose polymerase(PARP)proteins in the cells in various groups;imunofluorescence method was used to detect the expression levels of cleaved Caspase-3 protein in the cells in various groups.Fifteen model mice with subcutaneous tumor xenografts were randomly divided into blank group(0 mg·kg-1·d-1 D-limonene),low dose of D-limonene group(200 mg·kg-1·d-1 D-limonene),and high dose of D-limonene group(400 mg·kg-1·d-1 D-limonene),and there were 5 mice in each group.The inhibitory rates of the tumor in vitro in various groups were calculated;HE staining and immunohistochemical staining were used to observe the morphology of subcutaneous tumor tissue of the mice in various groups and the growth curves of the tumor were drawn;immunohistochemical assay was used to detect the positive expression rates of Ki67 protein in subcutaneous tumor tissue of the mice in various groups;TUNEL staining was used to detect the apoptosis of the tumor cells in various groups.Results:In control group,the cells were spindle-shaped,in good condition,growing closely and adherently,with normal organelles and cytoplasm.After treated for 48 h,the cells in 0.6 mmol·L-1 D-limonene group showed reduced volume,intact but more permeable cell membranes,shrunken cytoplasm,internal vacuole structures,and some fragments floating in the solution.The cells in 0.8 and 1.0 mmol·L-1 D-limonene groups exhibited significant apoptotic bodies and were in an apoptotic state.The CCK-8 results showed that compared with control group,the inhibitory rates of proliferation of the U87,LN229,and GL261 cells in 0.6,0.8,and 1.0 mmol·L-1 D-limonene groups were significantly increased(P<0.01),the inhibitory rates of proliferation of the U87 and GL261 cells were significantly increased(P<0.01).The clone formation assay results showed that compared with control group,the clone formation rates of the U87,LN229,and GL261 cells in 0.4,0.6,and 0.8 mmol·L-1 D-limonene groups were significantly decreased(P<0.05 or P<0.01).The AnnexinⅤ-FITC/PI results showed that compared with control group,after treated with D-limonene for 48 h,the apoptotic rates of the LN229 cells in 0.6,0.8,and 1.0 mmol·L-1 D-limonene groups were significantly increased(P<0.01).The Western blotting results showed that compared with control group,the expression levels of Bax proteins in the LN229 cells in 0.6,0.8,and 1.0 mmol·L-1 D-limonene groups were significantly increased(P<0.01),while the expression levels of AKT and Bcl-2 proteins were significantly decreased(P<0.01),the expression level of PARP protein in the LN229 cells in 0.8 and 1.0 mmol·L-1 D-limonene group was significanthy increased(P<0.01).The immunofluorescence results showed that compared with control group,the expression levels of cleaved Caspase-3 protein in the LN229 cells in 0.6,0.8,and 1.0 mmol·L-1 D-limonene groups were significantly increased(P<0.01).Compared with blank group,the tumor volumes of the mice in low and high doses of D-limonene groups were significantly decreased(P<0.01).Compared with blank group,the tumor weights of the mice in low and high doses of D-limonene groups were significantly decreased(P<0.05),and the inhitory rates of tumor were significantly increased(P<0.05).The tumor cells in blank group were diffusely distributed,with deepened nuclear staining and increased nucleocytoplasmic ratio;a large number of degenerated and necrotic tumor cells were observed in tumor tissue of the mice in low and high doses of D-limonene groups.Compared with blank group,the positive expression rates of Ki67 protein in tumor tissue of the mice in low and high doses of D-limonene groups were significantly decreased(P<0.01).Compared with blank group,the apoptotic rates of tumor cells of the mice in low and high doses of D-limonene groups were significantly increased(P<0.01).Conclusion:D-limonene has the inhibitory effect on the proliferation of the GBM cells;its mechanism may be related to the regulation of AKT protein expression and the activation of the Caspase-3 pathway to induce the apoptosis.

Objective To investigate whether activation of mitochondrial acetal dehydrogenase 2(ALDH2)alleviates hypoxic pulmonary hypertension by regulating the SIRT1/PGC-1α signaling pathway.Methods Thirty 8-week-old C57 BL/6 mice were randomized into control,hypoxia,and hypoxia+Alda-1(an ALDH2 activator)group(n=10),and the mice in the latter two groups,along with 10 ALDH2 knockout(ALDH2-/-)mice,were exposed to hypoxia(10%O2,90%N2)with or without daily intraperitoneal injection of Alda-1 for 4 weeks.The changes in right ventricular function and pressure(RVSP)of the mice were evaluated by echocardiography and right ventricular catheter test,and pulmonary artery pressure was estimated based on RVSP.Pulmonary vascular remodeling,right ventricular injury,myocardial α-SMA expression,distal pulmonary arteriole muscle normalization,right ventricular cross-sectional area,myocardial cell hypertrophy,and right cardiac hypertrophy index were assessed with HE staining,immunofluorescence staining and WGA staining,and the expressions of ALDH2,SIRT1,PGC-1α,P16INK4A and P21CIP1 were detected.In pulmonary artery smooth muscle cells with hypoxic exposure,the effect of Alda-1 and EX527 on cell senescence and protein expressions was evaluated using β-galactose staining and Western blotting.Results The wild-type mice with hypoxic exposure showed significantly increased RVSP,right ventricular free wall thickness and myocardial expressions of P16INK4A and P21CIP1,which were effectively lowered by treatment with Alda-1 but further increased in ALDH2-/-mice.In cultured pulmonary artery smooth muscle cells,hypoxic exposure significantly increased senescent cell percentage and cellular expressions of P16INK4A and P21CIP1,which were all lowered by treatment with Alda-1,but its effect was obviously attenuated by EX527 treatment.Conclusion ALDH2 alleviates hypoxia-induced senescence of pulmonary artery smooth muscle cells by upregulating the SIRT1/PGC-1α signaling pathway to alleviate pulmonary hypertension in mice.

Objective:To explore the genetic basis for a Chinese pedigree affected with co-morbid Ornithine carbamoyl transferase deficiency (OTCD) and MECP2 duplication syndrome.Methods:A proband who was admitted to the Neonatal Intensive Care Unit of Gansu Provincial Maternal and Child Health Care Hospital on December 19, 2017 was selected as the study subject. High-throughput sequencing and multiplex ligation-dependent probe amplification (MLPA) were carried out for her pedigree, and short tandem repeat-based linkage analysis and chromosome copy number variation sequencing (CNV-seq) were used for the prenatal diagnosis.Results:The proband, a 3-day-old female, was found to harbor heterozygous deletion of exons 7-9 of the OTC gene. Based on the guidelines from the American College of Medical Genetics and Genomics (ACMG), the variant was classified as likely pathogenic (PVS1+ PM2_Supporting+ PP4). The proband was diagnosed with OTCD, which was in keeping with her acute encephalopathy and metabolic abnormalities (manifesting as hyperammonemia, decreased blood citrulline, and increased urine orotic acid). Prenatal diagnosis was carried out for the subsequent pregnancy. The fetus did not harbor the exons 7-9 deletion of the OTC gene, but was found to carry a duplication in Xq28 region (which encompassed the whole region of MECP2 duplication syndrome) and was positive for the SRY sequence. The same duplication was also found in the proband and her mother. Considering the possible existence of X-chromosome inactivation, the proband was diagnosed with two X-linked recessive disorders including OTCD and MECP2 duplication syndrome, and the fetus was determined as a male affected with the MECP2 duplication syndrome. Conclusion:Discoveries of the pathogenic variants underlying the OTCD and MECP2 duplication syndrome have enabled clinical intervention, treatment, genetic counseling and prenatal diagnosis for this pedigree.

This study aims to prepare vitexin albumin nanoparticles(VT-BSA-NPs) to alleviate the low bioavailability of vitexin(VT) in vivo due to its poor water solubility. VT micro powders were prepared by the antisolvent crystallization method, and the morphology, size, and physicochemical properties of VT micro powders were studied. The results showed that the VT micro powder had a particle size of(187.13±7.15) nm, an approximate spherical morphology, and a uniform size distribution. Compared with VT, the chemical structure of VT micro powders has not changed. VT-BSA-NPs were prepared from VT micro powders by desolvation-crosslinking curing method. The preparation process was screened by single factor test and orthogonal test, and the quality evaluation of the optimal prescription particle size, PDI, Zeta potential, EE, and morphology was performed. The results showed that the average particle size of VT-BSA-NPs was(124.33±0.47) nm; the PDI was 0.184±0.012; the Zeta potential was(-48.83±2.20) mV, and the encapsulation rate was 83.43%±0.39%, all of which met the formulation-related requirements. The morphological results showed that the VT-BSA-NPs were approximately spherical in appearance, regular in shape, and without adhesion on the surface. In vitro release results showed a significantly reduced release rate of VT-BSA-NPs compared with VT, indicating a good sustained release effect. LC-MS/MS was used to establish an analytical method for in vivo analysis of VT and study the plasma pharmacokinetics of VT-BSA-NPs in rats. The results showed that the specificity of the analytical method was good, and the extraction recovery was more than 90%. Compared with VT and VT micro powders, VT-BSA-NPs could significantly increase AUC, MRT, and t_(1/2), which was beneficial to improve the bioavailability of VT.
Rats ; Animals ; Serum Albumin, Bovine/chemistry* ; Chromatography, Liquid ; Tandem Mass Spectrometry ; Nanoparticles/chemistry* ; Particle Size ; Drug Carriers/chemistry*

OBJECTIVE: To investigate efficacy between arthroscopic popliteal cyst drainage and arthroscopic popliteal cyst resection. METHODS: From January 2013 to June 2021, 54 patients with popliteal cyst (Rausching-Lindgren gradeⅠto Ⅲ) were treated with arthroscopic surgery. There were 24 males and 30 females. The age ranged from 44 to 72 years old, with a mean of (62.67±6.08) years old. The course of the disease ranged from 1 to 72 months, with a mean of(15±14) months. Twenty-four patients (group A) were underwent arthroscopic internal drainage of popliteal cyst. Thirty patients (group B) were underwent arthroscopic resection of popliteal cyst. Preoperative main symptoms included knee pain, swelling, walking pain, popliteal swelling, popliteal mass and so on. After 1, 3, 6 months and 1, 2 years of surgery, routine outpatient follow-up was conducted to observe and compare the surgical time, bleeding volume, preoperative and postoperative visual analog scale (VAS), knee Lysholm score, and complications between two groups. RESULTS: All incisions healed at one stage after operation. All 54 patients were followed up, and the duration ranged from 6 months to 2 years, with an average of (13.89±4.29) months. There was no intraoperative vascular or nerve injury. Operation time and intraoperative blood loss of the two groups:group A of (62.08±9.55) min and (8.00±1.69) ml, group B of (69.50±6.99) min and (8.70±2.00) ml. Popliteal pain, swelling, limitation of flexion and extension were significantly relieved after operation. VAS before and one month after operation between two groups:group A of 5.38±1.21 and 2.63±0.71, group B of 5.60±1.26 and 2.80±0.81. Lysholm scores of knee joint before and 6 months after operation:group A of 62.59±4.99 and 89.74±2.90, group B of 63.87±3.23 and 89.02±2.35. Knee joint function improved significantly in both groups. In group A, 4 cases had popliteal cyst at 3 months after operation, and 2 cases had small isolated cyst at 1 year after operation. There was no recurrence of cyst in group B. CONCLUSION The results between two arthroscopic treatments of popliteal cyst are satisfactory, and there is no significant difference in the amount of blood loss, safety, postoperative pain VAS score and knee function recovery. It is suggested that arthroscopic resection of the cyst wall should be performed when the technique is mature, especially for large cysts and septal cysts.
Female ; Male ; Humans ; Adult ; Middle Aged ; Aged ; Popliteal Cyst/surgery* ; Drainage ; Cysts ; Knee Joint/surgery* ; Pain