BACKGROUND:We have successfully established an animal model of small ear pig in southern Yunnan province with internal tension relieving technique combined with autologous Achilles tendon for anterior cruciate ligament reconstruction,and verified the stability and reliability of the model.However,whether internal tension relieving technique can promote the ligamentalization process of autologous Achilles tendon graft has not been studied. OBJECTIVE:To investigate the differences in the process of ligamentalization between conventional reconstruction and internal reduction reconstruction of the anterior cruciate ligament by gross view,histology and electron microscopy. METHODS:Thirty adult female small ear pigs in southern Yunnan province were selected.Anterior cruciate ligament reconstruction was performed on the left knee joint with the ipsilateral knee Achilles tendon(n=30 in the normal group),and anterior cruciate ligament reconstruction was performed on the right knee joint with the ipsilateral knee Achilles tendon combined with the internal relaxation and enhancement system(n=30 in the relaxation group).The autogenous right forelimb was used as the control group;the anterior cruciate ligament was exposed but not severed or surgically treated.At 12,24,and 48 weeks after surgery,10 animals were sacrificed,respectively.The left and right knee joint specimens were taken for gross morphological observation to evaluate the graft morphology.MAS score was used to evaluate the excellent and good rate of the ligament at each time point.Hematoxylin-eosin staining was used to evaluate the degree of ligament graft vascularization.Collagen fibers and nuclear morphology were observed,and nuclear morphology was scored.Ultrastructural remodeling was evaluated by scanning electron microscopy and transmission electron microscopy. RESULTS AND CONCLUSION:(1)The ligament healing shape of the relaxation group was better at various time points after surgery,and the excellent and good rate of MAS score was higher(P<0.05).Moreover,the relaxation group could obtain higher ligament vascularization score(P<0.05).(2)The arrangement of collagen bundles and fiber bundles in the two groups gradually tended to be orderly,and the transverse fiber connections between collagen gradually increased and thickened,suggesting that the strength and shape degree of the grafts were gradually improved,but the ligament remodeling in the relaxation group was always faster than that in the normal group at various time points after surgery.(3)The diameter,distribution density,and arrangement degree of collagen fibers in the relaxation group were better than those in the normal group at all time points,especially in the comparison of collagen fiber diameter between and within the relaxation group(P<0.05).

Immobilized enzyme-based enzyme electrode biosensors, characterized by high sensitivity and efficiency, strong specificity, and compact size, demonstrate broad application prospects in life science research, disease diagnosis and monitoring, etc. Immobilization of enzyme is a critical step in determining the performance (stability, sensitivity, and reproducibility) of the biosensors. Random immobilization (physical adsorption, covalent cross-linking, etc.) can easily bring about problems, such as decreased enzyme activity and relatively unstable immobilization. Whereas, directional immobilization utilizing amino acid residue mutation, affinity peptide fusion, or nucleotide-specific binding to restrict the orientation of the enzymes provides new possibilities to solve the problems caused by random immobilization. In this paper, the principles, advantages and disadvantages and the application progress of enzyme electrode biosensors of different directional immobilization strategies for enzyme molecular sensing elements by specific amino acids (lysine, histidine, cysteine, unnatural amino acid) with functional groups introduced based on site-specific mutation, affinity peptides (gold binding peptides, carbon binding peptides, carbohydrate binding domains) fused through genetic engineering, and specific binding between nucleotides and target enzymes (proteins) were reviewed, and the application fields, advantages and limitations of various immobilized enzyme interface characterization techniques were discussed, hoping to provide theoretical and technical guidance for the creation of high-performance enzyme sensing elements and the manufacture of enzyme electrode sensors.

Laparoscopic subtotal cholecystectomy (LSC) has been a safe and viable alternative to conversion to laparotomy in cases of severe cholecystitis. The objective of this study is to determine the utility of intraoperative choledochoscopy in LSC for the exploration of the gallbladder, cyst duct, and subsequent stone clearance of the cystic duct in cases of severe cholecystitis. A total of 72 patients diagnosed with severe cholecystitis received choledochoscopy-assisted laparoscopic subtotal cholecystectomy (CALSC). A choledochoscopy was performed to explore the gallbladder cavity and/or cystic duct, and to extract stones using a range of techniques. The clinical records, including the operative records and outcomes, were subjected to analysis. No LSC was converted to open surgery, and no bile duct or vascular injuries were sustained. All stones within the cystic duct were removed by a combination of techniques, including high-frequency needle knife electrotomy, basket, and electrohydraulic lithotripsy. A follow-up examination revealed the absence of residual bile duct stones, with the exception of one common bile duct stone, which was extracted via endoscopic retrograde cholangiopancreatography. In certain special cases, CALSC may prove to be an efficacious treatment for the management of severe cholecystitis. This technique allows for optimal comprehension of the situation within the gallbladder cavity and cystic duct, facilitating the removal of stones from the cystic duct and reducing the residue of the non-functional gallbladder remnant.

Laparoscopic subtotal cholecystectomy (LSC) has been a safe and viable alternative to conversion to laparotomy in cases of severe cholecystitis. The objective of this study is to determine the utility of intraoperative choledochoscopy in LSC for the exploration of the gallbladder, cyst duct, and subsequent stone clearance of the cystic duct in cases of severe cholecystitis. A total of 72 patients diagnosed with severe cholecystitis received choledochoscopy-assisted laparoscopic subtotal cholecystectomy (CALSC). A choledochoscopy was performed to explore the gallbladder cavity and/or cystic duct, and to extract stones using a range of techniques. The clinical records, including the operative records and outcomes, were subjected to analysis. No LSC was converted to open surgery, and no bile duct or vascular injuries were sustained. All stones within the cystic duct were removed by a combination of techniques, including high-frequency needle knife electrotomy, basket, and electrohydraulic lithotripsy. A follow-up examination revealed the absence of residual bile duct stones, with the exception of one common bile duct stone, which was extracted via endoscopic retrograde cholangiopancreatography. In certain special cases, CALSC may prove to be an efficacious treatment for the management of severe cholecystitis. This technique allows for optimal comprehension of the situation within the gallbladder cavity and cystic duct, facilitating the removal of stones from the cystic duct and reducing the residue of the non-functional gallbladder remnant.

Laparoscopic subtotal cholecystectomy (LSC) has been a safe and viable alternative to conversion to laparotomy in cases of severe cholecystitis. The objective of this study is to determine the utility of intraoperative choledochoscopy in LSC for the exploration of the gallbladder, cyst duct, and subsequent stone clearance of the cystic duct in cases of severe cholecystitis. A total of 72 patients diagnosed with severe cholecystitis received choledochoscopy-assisted laparoscopic subtotal cholecystectomy (CALSC). A choledochoscopy was performed to explore the gallbladder cavity and/or cystic duct, and to extract stones using a range of techniques. The clinical records, including the operative records and outcomes, were subjected to analysis. No LSC was converted to open surgery, and no bile duct or vascular injuries were sustained. All stones within the cystic duct were removed by a combination of techniques, including high-frequency needle knife electrotomy, basket, and electrohydraulic lithotripsy. A follow-up examination revealed the absence of residual bile duct stones, with the exception of one common bile duct stone, which was extracted via endoscopic retrograde cholangiopancreatography. In certain special cases, CALSC may prove to be an efficacious treatment for the management of severe cholecystitis. This technique allows for optimal comprehension of the situation within the gallbladder cavity and cystic duct, facilitating the removal of stones from the cystic duct and reducing the residue of the non-functional gallbladder remnant.

Palliative care is recognized as an effective measure to improve the quality of life for patients with end-stage diseases， and the significance and role of such patients participating in clinical trials to conquer major diseases has also become a broad consensus. However， due to the special physical， psychological， and social conditions of terminal trial participants， the ethical problems encountered in the trial process are more serious and complex. Drawing on ethical practice experience， these seemingly common phenomena and issues were deeply analyzed. Combined with the palliative care philosophy for end-stage patients， this paper proposed a series of improvement suggestions throughout the entire life cycle of clinical trials， hoping to promote the quality improvement of clinical research in which end-stage patients participate as subjects， while effectively protecting the safety and rights of the subjects and ensuring they receive appropriate palliative care during their participation in clinical trials or clinical-related scientific research.

Aim To investigate the effect of ellagic acid (EA) on cognitive function in APP/PS 1 double- transgenic mice, and to explore the regulatory mechanism of ellagic acid on the level of oxidative stress in the hippocampus of double-transgenic mice based on the phosphatidylinositol 3-kinase/protein kinase B/glycogen synthase kinase-3 (PI3K/AKT/GSK-3 β) signaling pathway. Methods Thirty-two SPF-grade 6-month-old APP/PS 1 double transgenic mice were randomly divided into four groups, namely, APP/PS 1 group, APP/PS1 + EA group, APP/PS1 + LY294002 group, APP/PS 1 + EA + LY294002 group, with eight mice in each group, and eight SPF-grade C57BL/6J wild type mice ( Wild type) were selected as the blank control group. The APP/PS 1 + EA group was given 50 mg · kg

Aim To investigate whether alisol A (AA) could improve the blood brain barrier (BBB) mediated cortex cerebral ischemia-repeifusion injury (CIRI) by inhibiting matrix metalloproteinase 9 (MMP-9). Methods The global cerebral ischemia- reperfusion (GCI/R) model in mice was established, and the AA was intragastric injected subsequently for seven days. The modified neurological severity scores (mNSS), open field test and Y-maze test were applied to detect neurological function. Magnetic resonance spectroscopy (MRS) was used to detect relevant neu- rosubstance metabolism in cortex of mice. Transmission electron microscope (TEM) was employed to observe the ultrastructure of BBB in cortex. Western blot and immunohistochemistry were used to detect the MMP-9 level in cortex. The binding possibility of A A and MMP-9 was determined by molecular docking. Results Compared with Sham group, mice in GCI/R group have an increased mNSS score but decreased at total distance and center distance to total distance ratio in open field test as well as alternation rate in Y-maze test (P<0.01). While mice in GCI/R + AA group have a decreased mNSS score but increased at total distance and center distance to total distance ratio in open field test as well as alternation rate in Y-maze test (P<0.01) compared with GCI/R group. MRS results found that in cortex of GCI/R group mice, the level of GABA and NAA significantly decreased while the Cho, mI and Tau level increased (P<0.01). Whereas in GCI/R + AA group mice, the GABA and NAA level increased and the Cho, ml and Tau decreased significantly (P<0.01). By TEM we observed that the basilemma of cerebral microvessels collapsed, the lumen narrowed, the endothelial cells were active and plasma membranes ruffled, gaps between cells were enlarged and tight junctions were damaged and the end feet of astrocytes were swollen in GCI/R group mice. While in GCI/R + AA group mice, the lumen was filled, plasma membranes of endothelial cells were smooth, tight junctions were complete and end feet of astrocytes were in normal condition. Western blot and immunohistochemistry both found that the MMP-9 level increased in GCI/R group mice (P < 0.01) and decreased in GCI/R + AA group mice (P < 0.05). Molecular docking proved the binding between aliso A and MMP9 through TYR-50 and ARG-106, and the binding energy was calculated as -6.24 kcal · mol

Aim To explore the effect of kaempferol-7- 0-neohesperidoside (K70N) against prostate cancer (PCa) and the underlying mechanism. Methods The effect of K70N on the proliferation of PCa cell lines PC3, DU145, C4-2 and LNCaP was detected using CCK8 assay. The effect of K70N on migration ability of DU145 cells was determined by wound healing assay. The targets of K70N and PCa were screened from SuperPred and other databases. The common targets both related to K70N and PCa were obtained from the Venny online platform, a protein-protein interaction network (PPI) was constructed by the String and Cyto- scape. Meanwhile, the GO and KEGG functional enrichment were analyzed by David database. Then, a "drug-target-disease-pathway" network model was constructed. Cell cycle of PCa cells treated with K70N was analyzed by flow cytometry. The expressions of cycle-associated proteins including Skp2, p27 and p21 protein were detected by Western blot. Molecular docking between Skp2 and K70N was conducted by Sybyl X2. 0. Results K70N significantly inhibited the proliferation and migration of PCa cells. A total number of 34 drug-disease intersection targets were screened. The String results showed that Skp2 and p27, among the common targets, were the key targets of K70N for PCa treatment. Furthermore, GO and KEGG functional en-richment indicated that the mechanism was mainly related to the cell cycle. Flow cytometry showed that K70N treatment induced cell cycle arrest at the S phase. Compared with the control group, the protein expression level of Skp2 was significantly down-regulated, while the protein expression levels of p27 and p21 were up-regulated. The network molecular docking indicated that the ligand K70N had a good binding ability with the receptor Skp2. Conclusions K70N could inhibit the proliferation and migration of PCa cells, block the cell cycle in the S phase, which may be related to the regulation of cell cycle through the Skp2- p27/p21 signaling pathway.

ObjectiveTo observe the intervention effect of Huaiqihuang granules （HQH） on immunoglobulin A vasculitis nephritis （IgAVN） mice and explore the underlying therapeutic mechanism. MethodFifty SPF-grade male Kunming mice were randomly divided into a normal group， an IgAVN model group， a dexamethasone group （2.5 mg·kg^-1·d^-1）， a low-dose HQH group （4 g·kg^-1·d^-1）， and a high-dose HQH group （8 g·kg^-1·d^-1）. The mouse model was established using oral administration of gliadin combined with intravenous injection of India ink. After successful modeling， the mice were euthanized after 4 weeks of gastric gavage according to groups. The 24 h urinary total protein （24 h UTP）， urine β₂-microglobulin （β₂-MG）， serum total protein， albumin， IgA， etc. were detected in each group. Flow cytometry was used to determine the proportion of T helper 17 （Th17） cells in spleen cell suspension. Western blot was employed to detect the expression of adenosine 5'-monophosphate-activated protein kinase α （AMPKα）， phosphorylated AMPKα （p-AMPKα）， acetyl-CoA carboxylase 1 （ACC1）， and phosphorylated ACC1 （p-ACC1） in Th17 cells. Pathological changes in the spleen and kidneys were observed. ResultCompared with the normal group， the IgAVN model group showed significant increases in 24 h UTP， urine β₂-MG， total cholesterol （P<0.05）， serum interleukin-17 （IL-17）， IgA， Th17 proportion in the spleen cell suspension， and IL-17 expression in the spleen tissue （P<0.01）， and significantly decreased serum total protein， albumin， p-AMPKα/AMPKα， and p-ACC1/ACC1 expression of Th17 cells （P<0.01）. Compared with the IgAVN model group， in the 4th week， the 24 h UTP， urine β₂-MG， serum IL-17， IgA levels， and renal IgA deposition were significantly reduced in each treatment group （P<0.01）， and the Th17 proportion and IL-17 expression in spleen tissue were significantly decreased （P<0.05， P<0.01）. Serum albumin levels significantly increased （P<0.05）. Compared with the IgAVN model group， the dexamethasone group and the high-dose HQH group showed increases in serum total protein （P<0.01）， p-AMPKα/AMPKα， and p-ACC1/ACC1 expression of Th17 cells （P<0.05， P<0.01）. The high-dose HQH group showed a significant decrease in total cholesterol level （P<0.05）. Various treatment groups showed different degrees of improvement in spleen and kidney pathological changes. ConclusionHQH may affect Th17 cell differentiation by regulating the AMPK/ACC pathway， correcting immune inflammatory disorders， and exerting therapeutic effects on IgAVN.