OBJECTIVE To focus on the classic NOD-like receptor protein 3 （NLRP3）/Caspase-1/gasdermin D （GSDMD） pyroptosis pathway and explore the mechanism by which Huatan qushi huoxue formula （HQHF） inhibits macrophage pyroptosis to ameliorate metabolic dysfunction-associated steatohepatitis （MASH）. METHODS RAW264.7 cells were divided into 5 groups： Control group （10% blank serum）， Model group [10% blank serum+5 μg/mL lipopolysaccharide （LPS）]， HQHF-L group （2.5% drug-containing serum+7.5% blank serum+5 μg/mL LPS）， HQHF-M group （5% drug-containing serum+5% blank serum+5 μg/mL LPS）， and HQHF-H group （10% drug-containing serum+5 μg/mL LPS）. After 24 h of routine culture post-administration， cells and supernatants were collected for assays. Cell morphology was observed via scanning electron microscopy and phase-contrast microscopy； localization and expression of gasdermin D-N （GSDMD-N） were observed by immunofluorescence. Interleukin-1β （IL-1β） and IL-18 contents in supernatants were detected by ELISA； mRNA and protein expressions of NLRP3， Caspase-1， and GSDMD were measured using real-time PCR and Western blot. RESULTS Compared with the Control group， the Model group showed typical pyroptotic morphology （cell membrane bulging and pore formation）， increased aggregation and fluorescence intensity of GSDMD-N on the cell membrane （ P ＜0.05）， significantly increased the contents of IL-1β and IL-18 in cell supernatants （ P ＜0.05）， and significantly up-regulated mRNA and protein expressions of NLRP3， Caspase-1， and GSDMD in cells （ P ＜0.05）. Compared with the Model group， the HQHF-L， HQHF-M and HQHF-H groups showed improved pyroptotic morphology， reduced membrane localization and significantly weakened fluorescence intensity of GSDMD-N （ P ＜0.05）， significantly decreased the contents of IL-1β and IL-18 in cell supernatants （ P ＜0.05）， and significantly down-regulated mRNA and protein expressions of NLRP3， Caspase-1， and GSDMD in cells （ P ＜0.05）. CONCLUSIONS HQHF inhibits LPS-induced macrophage pyroptosis， and its mechanism of improving MASH may be associated with the suppression of the activation of the classical NLRP3/Caspase-1/GSDMD pyroptosis pathway.

Cemental tear is a rare and indetectable condition unless obvious clinical signs present with the involvement of surrounding periodontal and periapical tissues. Due to its clinical manifestations similar to common dental issues, such as vertical root fracture, primary endodontic diseases, and periodontal diseases, as well as the low awareness of cemental tear for clinicians, misdiagnosis often occurs. The critical principle for cemental tear treatment is to remove torn fragments, and overlooking fragments leads to futile therapy, which could deteriorate the conditions of the affected teeth. Therefore, accurate diagnosis and subsequent appropriate interventions are vital for managing cemental tear. Novel diagnostic tools, including cone-beam computed tomography (CBCT), microscopes, and enamel matrix derivatives, have improved early detection and management, enhancing tooth retention. The implementation of standardized diagnostic criteria and treatment protocols, combined with improved clinical awareness among dental professionals, serves to mitigate risks of diagnostic errors and suboptimal therapeutic interventions. This expert consensus reviewed the epidemiology, pathogenesis, potential predisposing factors, clinical manifestations, diagnosis, differential diagnosis, treatment, and prognosis of cemental tear, aiming to provide a clinical guideline and facilitate clinicians to have a better understanding of cemental tear.
Humans ; Dental Cementum/injuries* ; Consensus ; Diagnosis, Differential ; Cone-Beam Computed Tomography ; Tooth Fractures/therapy*

Successful cloning through somatic cell nuclear transfer (SCNT) faces significant challenges due to epigenetic obstacles. Recent studies have highlighted the roles of H3K4me3 and H3K27me3 as potential contributors to these obstacles. However, the underlying mechanisms remain largely unclear. In this study, we generated genome-wide maps of H3K4me3 and H3K27me3 in mouse pre-implantation NT embryos. Our analysis revealed that aberrantly over-represented broad H3K4me3 domain and H3K27me3 signal lead to increased bivalent marks at gene promoters in NT embryos compared with naturally fertilized (NF) embryos at the 2-cell stage, which may link to relatively low levels of H3K36me3 in NT 2-cell embryos. Notably, the overexpression of Setd2, a H3K36me3 methyltransferase, successfully restored multiple epigenetic marks, including H3K36me3, H3K4me3, and H3K27me3. In addition, it reinstated the expression levels of ZGA-related genes by reestablishing H3K36me3 at gene body regions, which excluded H3K27me3 from bivalent promoters, ultimately improving cloning efficiency. These findings highlight the excessive bivalent state at gene promoters as a potent barrier and emphasize the removal of these barriers as a promising approach for achieving higher cloning efficiency.
Animals ; Mice ; Histone-Lysine N-Methyltransferase/biosynthesis* ; Histones/genetics* ; Nuclear Transfer Techniques ; Female ; Gene Expression Regulation, Developmental ; Promoter Regions, Genetic ; Epigenesis, Genetic ; Embryo, Mammalian/metabolism*

This study aimed to investigate the anti-inflammatory and antioxidant effects of atractylon on lipopolysaccharide (LPS)-induced acute lung injury (ALI) in mice. Changes in lung function parameters were measured in mice after intraperitoneal administration of atractylon. Pathological changes in lung tissue were observed by H&E staining, and the degree of pulmonary edema was assessed by the lung wet/dry weight ratio (W/D). Kit assays were used to detect changes in oxidative stress markers in mouse serum and the protein concentration in bronchoalveolar lavage fluid (BALF). ELISA was employed to measure the expression levels of inflammatory cytokines in BALF and serum. Western blot was used to detect the expression levels of proteins related to the cGAS-STING pathway and vascular cell adhesion molecule-1 (VCAM-1) in lung tissue. Results showed that, compared to the ALI model group, mice in the low-dose and high-dose atractylon groups exhibited significant improvement in lung function parameters, alleviated pulmonary edema, and reduced inflammatory cell infiltration in lung tissue. Protein content and inflammatory cytokine levels in serum and BALF were decreased, while serum oxidative stress indicators were improved. Western blot results further indicated that atractylon could regulate the cGAS-STING pathway, blocking the generation of inflammatory signals, and simultaneously inhibit VCAM-1 expression, thereby reducing pulmonary vascular injury. The results suggest that atractylon may alleviate LPS-induced ALI by modulating the cGAS-STING signaling pathway, reducing the expression of pro-inflammatory cytokines and the production of pro-inflammatory mediators, and improving vascular endothelial injury. This study provides a new potential target and theoretical basis for the treatment of ALI, as well as a potential drug candidate for ALI therapy.

Congenital portosystemic shunts（CPSS）are vascular malformations caused by developmental anomalies that create abnormal shunts between the portal and systemic veins.This anomaly permits partial or complete bypass of portal blood flow from the liver，leading it directly into the systemic circulation without hepatic filtration.If untreated，CPSS can lead to multisystem complications，with pulmonary vascular complications being the most severe outcome.These complications are complex in mechanism，often insidious in onset，and present with nonspecific symptoms，increasing the risk of delayed diagnosis and significantly impacting the quality of children lives.Pulmonary vascular complications associated with CPSS primarily include portopulmonary hypertension，characterized by pulmonary vascular remodeling，and hepatopulmonary syndrome，distinguished by intrapulmonary vascular dilatation and abnormal arterial oxygenation.This article aims to review the anatomical classifications of CPSS，along with the pathophysiology，clinical presentation，diagnosis，and treatment of its pulmonary vascular complications，with the goal of enhancing clinical awareness and providing a reference for diagnosis and management.

Objective To observe the influence of the staining phenomenon after fluorescein sodium staining on eye irritation in normal rabbits.MethodsIn the experimental rabbit eye irritation test conducted with sodium chloride eye drops, Siwei Zhenceng Bingpeng eye drops, sodium hyaluronate eye drops, sodium cromoglycate eye drops, and compound aspartate eye drops (4 in each group, half male and half female), the left eyes of rabbits were administered normal saline (self-negative control) and the right eyes were administered the experimental medicine; the eyes were stained with 1% sodium fluorescein, and eye irritation was observed and scored using slit lamp microscope for 31 days. Morphological changes of corneal epithelial staining were recorded and the incidence of staining was calculated. After the observation, the eyeballs and Hasselblad glands were examined histopathologically, and the staining rate of the left eye was compared with that of the right eye which was administered the corresponding medicine.ResultsNeither eye had any irritation symptoms; the scores were 0, and the total incidences of corneal staining were 3% (left) and 1% (right), respectively. There was no significant difference between the two groups (P > 0.05). Corneal epithelial staining showed single-spot staining, scattered dot, localized, or large areas of fusion staining. No histopathological changes were found in the eyeballs or Hasselblad glands, and the results were evaluated as non-irritative.Conclusion The irregularity of corneal epithelial staining in rabbits did not influence the results of the ocular irritation test.

Abstract： Objective　To elucidate the antigenic antibody reaction of recombinant expression of non-structural protein 1 (NS1) of Japanese encephalitis (JE) virus with various mosquito-borne flaviviruses, including JE virus, and the antigenic antibody reaction of serum samples of patients infected with JE virus in acute stage. Methods　In this study, Escherichia coli prokaryotic expression vector (pET) system was used to recombinant express Japanese encephalitis virus NS1 gene. Western Blot assay was performed to detect the antibody responses of the recombinantly expressed protein against a variety of mosquito-transmitted flaviviruses, including JE virus, as well as antigen-antibody reactions of serum from patients with acute JE virus infection. Results　The NS1 gene expression product of JE virus (P3 strain) was in the form of an inclusion body, and the denatured and renatured expression product was displayed as a single band in the denatured gel (polyacrylamide gel electrophoresis, PAGE), with a molecular weight of about 45 000. The results of further antigen-antibody analysis showed that the antigen/antibody hybridization reaction of the expression product with polyclonal or monoclonal antibody of JE virus (mosquito isolates, encephalitis isolates) and serum samples of patients with acute JE virus infection could be completely consistent. The recombinant product showed negative antigen/antibody hybridization reactions with mosquito-transmitted flaviviruses, such as dengue virus and yellow fever virus polyclonal antibodies, but positive reactions with polyclonal antibodies to West Nile virus and Murray Valley encephalitis virus. Conclusions　In this study, the recombinant expression of the NS1 protein of JE virus was successfully obtained, and the antigen/antibody reaction between the recombinant protein and samples of patients infected with mosquito-borne flavivirus and JE virus was analyzed. The study results provide important basic data for elucidating the antigen-antibody reaction between the NS1 protein of JE virus and mosquito-borne flavivirus. The recombinant expression protein obtained in this study provides an important material basis for further research on the function of JE virus NS1 protein.

Objective:To summarize the experience of Bipartition osteotomy combined with craniofacial bone remodeling in the treatment of orbital hypertelorism and midfacial dysplasia.Methods:A retrospective analysis was done by the clinical data of two patients with orbital hypertelorism with maxillofacial malformation treated in the Shanghai Ninth People’s Hospital Affiliated to Shanghai Jiao Tong University School of Medicine in July 2005 and August 2018. The Bipartition osteotomy via intracranial approach were applied for the two patients, one male and one female. The two patients aged 17 and 19, were suffered from degree Ⅰ orbital hypertelorism [interorbital distance(IOD) 32 mm and 34 mm]. With a frontal bone fenestration, the Monobloc osteotomy was done firstly to dislocate the cranio-orbital-maxilla bone graft, then the Bipartition osteotomy was performed according to the preoperative design through 3D computer aided design(CAD) and a V-shape bone graft was removed from the interorbital bone graft to split the hard palate longitudinally along the midline. After all, the whole facial bone was separated to two blocks: bilateral orbital-maxillary segments. Finally the midface had been remodeled by the bilateral orbital-maxillary segments which rotated and fixed internally. Self-rib nasal augmentation was done later. Patents’ complications, eye movement, visual acuity, olfactory sensation, nasal shape, IOD were measured through CT scan and the appearances were observed in the postoperative follow-up to determine the degree of improvement.Results:15 mm in width of interorbital bone was resected in both of the two patients, respectively. Postoperative IOD reduced by 17 mm, 19 mm, respectively. Mild cerebrospinal fluid leakage occurred in both patients after operation. They recovered after 5, 8 days of pillow-free horizontal position, respectively. The male patient developed local skin infection at the coronal incision and recovered after dressing change for 1 week. One week after operation, the female patient’s nose tip was partially broken near the nasal columella, and recovered after debridement and repairment. Follow up for 4-11 months showed that the eye movement, visual acuity, normal convergence and olfactory sensation were normal and no diplopia of 2 patients. The nasal appearances and orbital hypertelorisum of them were corrected obviously by follow-up after 4 months, but the nasal morphology and epicanthus still need further improvement.Conclusions:The Bipartition osteotomy can effectively treat the orbital hypertelorisum through the internal rotation fixation of bilateral orbital-maxillary segments, that makes the high arch palate lower and the occlusal plane of the upper jaw flattened.

Objective:To summarize the experience of Bipartition osteotomy combined with craniofacial bone remodeling in the treatment of orbital hypertelorism and midfacial dysplasia.Methods:A retrospective analysis was done by the clinical data of two patients with orbital hypertelorism with maxillofacial malformation treated in the Shanghai Ninth People’s Hospital Affiliated to Shanghai Jiao Tong University School of Medicine in July 2005 and August 2018. The Bipartition osteotomy via intracranial approach were applied for the two patients, one male and one female. The two patients aged 17 and 19, were suffered from degree Ⅰ orbital hypertelorism [interorbital distance(IOD) 32 mm and 34 mm]. With a frontal bone fenestration, the Monobloc osteotomy was done firstly to dislocate the cranio-orbital-maxilla bone graft, then the Bipartition osteotomy was performed according to the preoperative design through 3D computer aided design(CAD) and a V-shape bone graft was removed from the interorbital bone graft to split the hard palate longitudinally along the midline. After all, the whole facial bone was separated to two blocks: bilateral orbital-maxillary segments. Finally the midface had been remodeled by the bilateral orbital-maxillary segments which rotated and fixed internally. Self-rib nasal augmentation was done later. Patents’ complications, eye movement, visual acuity, olfactory sensation, nasal shape, IOD were measured through CT scan and the appearances were observed in the postoperative follow-up to determine the degree of improvement.Results:15 mm in width of interorbital bone was resected in both of the two patients, respectively. Postoperative IOD reduced by 17 mm, 19 mm, respectively. Mild cerebrospinal fluid leakage occurred in both patients after operation. They recovered after 5, 8 days of pillow-free horizontal position, respectively. The male patient developed local skin infection at the coronal incision and recovered after dressing change for 1 week. One week after operation, the female patient’s nose tip was partially broken near the nasal columella, and recovered after debridement and repairment. Follow up for 4-11 months showed that the eye movement, visual acuity, normal convergence and olfactory sensation were normal and no diplopia of 2 patients. The nasal appearances and orbital hypertelorisum of them were corrected obviously by follow-up after 4 months, but the nasal morphology and epicanthus still need further improvement.Conclusions:The Bipartition osteotomy can effectively treat the orbital hypertelorisum through the internal rotation fixation of bilateral orbital-maxillary segments, that makes the high arch palate lower and the occlusal plane of the upper jaw flattened.

Objective:To identify the gene mutations associated with facial cleft-related orbital hypertelorism in 3 pairs of monozygotic twins with different phenotypes (with/without hypertelorism) and to investigate their mechanisms.Methods:From May 2014 to May 2019, 3 pairs of monozygotic twins, 2 males and 4 females, aged 5-18 years, were treated in Ninth People’s Hospital Affiliated to Shanghai Jiao Tong University School of Medicine, one with normal orbital distance and widening of orbital distance was caused by facial fissure. Among the twins, there was 1 case of orbital hypertelorism and the other case of without orbital hypertelorism, and the hypertelorism was caused by facial cleft. To screen for mutations in hypertelorism, whole genome sequencing was performed on 3 pairs of twins. The Sanger method was used to sequence the exons of 33 patients with facial fissure associated hypertelorism and 50 healthy individuals in the same period to identify the genes selected by the whole genome sequencing. The periosteal tissues were obtained from patients and healthy people during plastic surgery. The cells were cultured, the activity of alkaline phosphatase was measured, and the osteogenic differentiation was identified by alizarin red staining, real-time quantitative PCR and Western blotting were used to detect the mRNA and protein expression of signal transduction pathways in periosteal cells.Results:Whole genome sequencing analysis showed that in all three sets of twins, a new synonymous mutation (c.1479G>A, p. Q493Q) was found in the MAML3. In Sanger exon sequencing, 17(51.5%) of 33 patients with hypertelorism carried the mutation, while no mutation was detected in 50 normal controls. The result of periosteum-derived cytology showed that the expression of MAML3 mRNA and protein in the patient-derived cells was lower than that in the healthy-derived cells. Three, 7, 14 days after osteoinduction, the ALP activity in the cells from the patients was higher than that from the healthy subjects (8.540±1.450, 20.740±2.514, 24.090±3.213 vs. 5.268±0.482, 11.680±1.527, 13.200±0.592; all P<0.05). Fourteen days after osteoinduction, the result of alizarin red staining showed that there were more erythema formation in the cells from the patients than those from the healthy subjects, these result suggest that MAML3 mutation may lead to over-differentiation of human periosteal-derived cells. The mRNA and protein expression levels of hes1 and hes5 downstream of the Notch signal pathway were down-regulated in the periosteal cells of the patients, while Wnt3a and β-catenin mRNA and protein expression levels were up-regulated in the Wnt signal pathway. Conclusions:The MAML3 gene (c.1479G>A, p. Q493Q) mutation is one of the causative genes of facial cleft-related hypertelorism. Notch and Wnt/β-catenin signaling pathway play an important role in the pathogenesis of hypertelorism.