Serine protease inhibitor Kazal-type (SPINK) is a skin keratinizing protease inhibitor, which was initially found in animal serum and is widely present in plants, animals, bacteria, and viruses, and they act as key regulators of skin keratinizing proteases and are involved in the regulation of keratinocyte proliferation and inflammation, primarily through the inhibition of deregulated tissue kinin-releasing enzymes (KLKs) in skin response. This process plays a crucial role in alleviating various skin problems caused by hyperkeratinization and inflammation, and can greatly improve the overall condition of the skin. Specifically, the different members of the SPINK family, such as SPINK5, SPINK6, SPINK7, and SPINK9, each have unique biological functions and mechanisms of action. The existence of these members demonstrates the diversity and complexity of skin health and disease. First, SPINK5 mutations are closely associated with the development of various skin diseases, such as Netherton’s syndrome and atopic dermatitis, and SPINK5 is able to inhibit the activation of the STAT3 signaling pathway, thereby effectively preventing the metastasis of melanoma cells, which is important in preventing the invasion and migration of malignant tumors. Secondly, SPINK6 is mainly distributed in the epidermis and contains lysine and glutamate residues, which can act as a substrate for epidermal transglutaminase to maintain the normal structure and function of the skin. In addition, SPINK6 can activate the intracellular ERK1/2 and AKT signaling pathways through the activation of epidermal growth factor receptor and protease receptor-2 (EphA2), which can promote the migration of melanoma cells, and SPINK6 further deepens its role in stimulating the migration of malignant tumor cells by inhibiting the activation of STAT3 signaling pathway. This process further deepens its potential impact in stimulating tumor invasive migration. Furthermore, SPINK7 plays a role in the pathology of some inflammatory skin diseases, and is likely to be an important factor contributing to the exacerbation of skin diseases by promoting aberrant proliferation of keratinocytes and local inflammatory responses. Finally, SPINK9 can induce cell migration and promote skin wound healing by activating purinergic receptor 2 (P2R) to induce phosphorylation of epidermal growth factor and further activating the downstream ERK1/2 signaling pathway. In addition, SPINK9 also plays an antimicrobial role, preventing the interference of some pathogenic microorganisms. Taken as a whole, some members of the SPINK family may be potential targets for the treatment of dermatological disorders by regulating multiple biological processes such as keratinization metabolism and immuno-inflammatory processes in the skin. The development of drugs such as small molecule inhibitors and monoclonal antibodies has great potential for the treatment of dermatologic diseases, and future research on SPINK will help to gain a deeper understanding of the physiopathologic processes of the skin. Through its functions and regulatory mechanisms, the formation and maintenance of the skin barrier and the occurrence and development of inflammatory responses can be better understood, which will provide novel ideas and methods for the prevention and treatment of skin diseases.

Objective: To explore the possibility of performing allogeneic platelet-rich plasma (PRP) treatment for patients who were not suitable for autologous PRP collection through case reports of two patients with refractory wounds treated with allogeneic PRP. Methods: The ABO-compatible allogeneic whole blood was centrifuged 3 times to obtain allogeneic PRP within 6 hours of blood collection. Then the qualified allogeneic PRP was applied to 2 cases of refractory wound on the same day. Results: The platelet concentration in allogeneic PRP was higher than 1 000×10 /L, and the test results of infectious diseases, as well as the mixing of red blood cells and white blood cells, met the standard of quality control. Both patients achieved satisfactory wound healing outcomes (3 d). Conclusions: For patients who were not suitable for autologous PRP treatment, allogeneic PRP might be a new option.

Objective To observe the regulating effect and mechanism of Yichang Sanjie Granules on intestinal flora and immune function in mice with colon cancer.Methods Sixty mice were randomly divided into six groups,i.e.,the normal group,the model group,the low-,medium-and high-dose groups of Yichang Sanjie Granules,and the overexpression of melanoma absent gene 2(AIM2)plasmid(pcDNA-AIM2)intervention group,with 10 mice in each group.Colorectal cancer model was prepared by oxidized azomethine(AOM)/dextran sulfate sodium(DSS)induction method in all groups except normal group.After drug administration,the survival curves of mice in each group were plotted and the tumor volume was calculated;serum levels of immunoglobulin(Ig)G,IgM,interleukin(IL)-1β and IL-18 were detected by enzyme-linked immunosorbent assay(ELISA);peripheral blood levels of CD3+,CD4+,CD8+ T cells were detected by flow cytometry;the splenic index was determined;Hematoxylin-eosin(HE)staining was used to observe the pathological changes in colon tissues;16S-rDNA intestinal flora sequencing was used to detect the α-diversity of intestinal flora and the structure of intestinal flora communities;and protein immunoblotting(Wetsern Blot)was used to detect the protein expressions of AIM2,apoptosis-associated speckled-like protein containing a CARD(ASC),and cystatinase-1(caspase-1)in colon tissues.Results Compared with the normal group,the survival rate,serum levels of IgG and IgM,peripheral blood levels of CD3+ and CD4+ and CD4+/CD8+ ratio,protein expression levels of colon tissue AIM2,ASC and caspase-1 in the model group were significantly decreased,and the tumor volume,serum levels of IL-1β and IL-18,peripheral blood level of CD8+,and splenic index were significantly increased(all P＜0.05),and the HE staining results showed the characteristic manifestations of colon cancer;compared with the model group,the survival rate,serum levels of IgG and IgM,peripheral blood levels of CD3+ and CD4+ and CD4+/CD8+ ratio,protein expression levels of colon tissue AIM2,ASC and caspase-1 in the low-,medium-and high-dose groups of Yichang Sanjie Granules and the pcDNA-AIM2 group were significantly increased,and the tumor volume,serum levels of IL-1β and IL-18,level of peripheral blood CD8+,and splenic index were significantly decreased(all P＜0.05),and the HE staining results showed the manifestations of colon cancer were improved.Compared with the normal group,the Observed index,Chao1 index,Shannon index,the relative abundance of Bacteroidetes,Proteobacteria,Muribaculaceae,Lachnospiraceae-NK4A136group,and Ruminiclostridium in the model group were significantly decreased,while the relative abundance of Firmicutes,Actinobacteria,Patescibateria,Lactobacillus,Odoribacter,Alistipes,Ruminococcaceae-uncultured and Bacteroides was increased in the model group(P＜0.05);compared with the model group,the Observed index,Chao1 index,Shannon index,the relative abundance of Bacteroidetes,Proteobacteria,Muribaculaceae,Lachnospiraceae-NK4A136group and Ruminiclostridium were significantly increased,and the relative abundance of Firmicutes,Actinobacteria,Patescibateria,Lactobacillus,Odoribacter,Alistipes,Ruminococcaceae-uncultured and Bacteroides was decreased in the low-,medium-and high-dose groups of Yichang Sanjie Granules and the pcDNA-AIM2 group(all P＜0.05).Conclusion Yichang Sanjie Granules can increase autoimmunity and improve intestinal flora structure in mice with colon cancer,and its mechanism is related to the activation of AIM2 inflammatory vesicles.

Objective To enhance the standardization of surgical operations,shorten the learning curve,and reduce surgical complications by summarizing common errors and proposing solutions for beginners during pre-study of cavity construction in non-inflatable axillary approach laparoscopic thyroid surgery.Methods A retrospective analysis was made on 210 patients who underwent noninflatable axillary approach luminal thyroid surgery at the Department of Head and Neck Surgery,Zhejiang Provincial People's Hospital,between January 2022 and June 2023.Among them,150 patients were luminalized by the practitioners in the practice group,while 60 patients were luminalized by the practitioners in the skilled group.The occurrence of nine common errors during cavity construction,such as supraclavicular nerve injury and scapulohumeral muscle injury,was assessed and compared between the two groups.Results There was no statistically significant difference in age,gender,tumor diameter,tumor location or pathology type between the two groups(P＞0.05).The staff in the practice group had a significant difference in supraclavicular cutaneous nerve injury(19.33％vs.3.33％),scapulohumeral muscle injury or disarticulation(16.00％vs.0),anterior cervical band of free excess(24.00％vs.3.33％),pectoralis major muscle and supraclavicular fascia injury(16.00％vs.5.00％),external jugular vein and genuine branch injury(8.00％vs.0),excessive freeing of the sternoclavicular joint(7.33％vs.0),and incorrect entry of the sternocleidomastoid muscle gap(8.67％vs.0)were significantly higher in incidence than the employees in the skilled group(P＜0.05).There was no statistically significant difference between employees in the practice group and those in the skilled group in terms of injury to the internal jugular vein and its geniculate branches(4.00％vs.0),and incorrect entry of the anterior cervical strap muscles into the hierarchy(2.00％vs.0)(P＞0.05).Conclusion The incidence of damage to the surrounding tissue was significantly higher in the novice group than in the expert group during endoscopic thyroid surgery for space construction.A solid anatomical foundation and proficient endoscopic surgical technique serve as fundamental prerequisites for achieving a successful space system construction.By summarizing common challenges and providing corresponding solutions encountered during endoscopic thyroid surgery,this study further enhances and refines the learning system for total endoscopic thyroid procedures.

Objective:To evaluate the value of 18F-FDG PET/CT for preoperative localization of epileptogenic foci in refractory epilepsy patients with negative MRI. Methods:Clinical data (550 lobes) of 55 epilepsy patients (38 males, 17 females, age (20.0±8.1) years) with negative MRI who underwent preoperative 18F-FDG PET/CT-MRI between January 2014 and June 2020 at the First Affiliated Hospital of Jinan University were retrospectively analyzed. The sensitivity, specificity, accuracy, positive predictive value (PPV) and negative predictive value (NPV) of 18F-FDG PET/CT, video electroencephalogram (VEEG), PET/CT+ VEEG and PET/CT-VEEG for localizing epileptogenic foci were calculated using stereoelectroencephalography (SEEG) and the outcomes of at least 1 year of postoperative follow-up as reference standards. χ2 test was used to compare the efficiencies of different examination modalities for unilobar, multilobar and all patients. Results:The correct lateralization rate of epileptogenic foci was 92.6%(25/27) using PET/CT. The sensitivity, specificity, accuracy, PPV and NPV of PET/CT for localization of epileptogenic foci were 65.1%(54/83), 77.9%(364/467), 76.0%(418/550), 34.4%(54/157) and 92.6%(364/393), respectively. The sensitivities of PET/CT-VEEG for localization of epileptogenic foci in all patients and patients with multilobar epilepsy were higher than those of VEEG alone (75.9%(63/83) vs 45.8%(38/83), 68.6%(35/51) vs 31.4%(16/51); χ2 values: 15.80, 14.16, both P<0.001). The specificities of PET/CT+ VEEG for localization of epileptogenic foci in all patients and patients with unilobar epilepsy were higher than those of VEEG alone (97.6%(456/467) vs 94.6%(442/467), 97.9%(282/288) vs 94.1%(271/288); χ2 values: 5.66, 5.48; P values: 0.017, 0.019). The sensitivity of PET/CT-VEEG (PET/CT and VEEG concordance) for localization of epileptogenic foci was higher than that of PET/CT+ VEEG (PET/CT and VEEG discordance) (8/9 vs 28.4%(21/74); χ2=10.40, P=0.001), and its specificity and accuracy were higher than those of PET/CT-VEEG (PET/CT and VEEG discordance) (93.4%(57/61) vs 71.7%(291/406), 92.9%(65/70) vs 72.1%(346/480); χ2 values: 13.23, 13.96; both P<0.001). Conclusions:18F-FDG PET/CT can localize and lateralize epileptogenic foci in patients with negative MRI. The combination of 18F-FDG PET/CT and VEEG improves the sensitivity, specificity, and accuracy for epileptogenic foci detection. 18F-FDG PET/CT is more accurate in detecting epileptogenic foci when it is concordant with VEEG.

OBJECTIVE To investigate the effect of leech freeze-dried powder combined with scorpion freeze-dried powder on carotid artery thrombosis in rats. METHODS FeCl3 was used to induce carotid artery thrombosis, and the experiment was divided into 6 groups: sham-operated group(sham group), model group, leech freeze-dried powder group(leech group), whole scorpion freeze-dried powder group(scorpion group), leech+scorpion half-dose group(half-dose group), leech+scorpion full-dose group(full-dose group), 6 rats in each group. The inhibition rate of bilateral carotid artery thrombosis was calculated after the final administration. The activated partial thromboplastin time(APTT), plasma prothrombintime(PT), plasma fibrinogen(FIB), and prothrombin time(TT) were measured by platelet aggregation and coagulation factor analyzer. The levels of plasma tissue-type plasminogen activator(t-PA) and plasminogen activatorinhibitor-I(PAI-I) were measured by ELSIA, and the thrombosis of each group was observed by HE staining. RESULTS Compared with model group, the wet weight and dry weight of thrombus were reduced in the leech, scorpion, half-dose and full-dose groups, and the full-dose group had the best effect in inhibiting thrombus formation. Compared with sham group, APTT, PT, TT was shortened, FIB content increased and t-PA level decreased in model group; compared with model group, APTT, PT, TT was prolonged, FIB content decreased and t-PA level increased in each dose group. The PAI-Ⅰ was not significantly different in each group. A large number of thrombus were seen in the blood vessels of model group. Compared with model group, there were significantly fewer thrombus in the leech, scorpion and half-dose groups, and almost no significant thrombus in the full-dose group. CONCLUSION Leech freeze-dried powder combined with scorpion freeze-dried powder inhibited carotid artery thrombosis in rats by affecting coagulation function and fibrinolytic system.

Objective To investigate the role of RNA-binding motif protein X-linked(RBMX)in regulating the proliferation,migration,invasion and glycolysis in human bladder cancer cells.Methods A lentivirus vectors system and RNA interference technique were used to construct bladder cancer 1376 and UC-3 cell models with RBMX overexpression and knockdown,respectively,and successful cell modeling was verified using RT-qPCR and Western blotting.Proliferation and colony forming ability of the cells were evaluated using EdU assay and colony-forming assay,and cell migration and invasion abilities were determined using Transwell experiment.The expressions of glycolysis-related proteins M1 pyruvate kinase(PKM1)and M2 pyruvate kinase(PKM2)were detected using Western blotting.The effects of RBMX overexpression and knockdown on glycolysis in the bladder cancer cells were assessed using glucose and lactic acid detection kits.Results RT-qPCR and Western blotting confirmed successful construction of 1376 and UC-3 cell models with RBMX overexpression and knockdown.RBMX overexpression significantly inhibited the proliferation,clone formation,migration and invasion of bladder cancer cells,while RBMX knockdown produced the opposite effects.Western blotting results showed that RBMX overexpression increased the expression of PKM1 and decreased the expression of PKM2,while RBMX knockdown produced the opposite effects.Glucose consumption and lactate production levels were significantly lowered in the cells with RBMX overexpression(P<0.05)but increased significantly following RBMX knockdown(P<0.05).Conclusion RBMX overexpression inhibits bladder cancer progression and lowers glycolysis level in bladder cancer cells by downregulating PKM2 expression,suggesting the potential of RBMX as a molecular target for diagnosis and treatment of bladder cancer.

ObjectiveTo investigate the effects of berbamine hydrochloride on sorafenib resistance in hepatocellular carcinoma cells and the underlying mechanisms. MethodThe sorafenib-resistant cell line SMMC-7721/S was selected by the concentration increment method starting at 1.25 μmol·L^-1 sorafenib. Both SMMC-7721 and SMMC-7721/S cells were treated with 0， 2.5， 5， 10， 15， 20 μmol·L^-1 sorafenib， and the cell counting kit-8 （CCK-8） assay was employed to determine the half maximal inhibitory concentration （IC₅₀） and calculate the resistance index （RI）. Western blot was conducted to compare the expression of proteins involved in autophagy and phosphatidylinositol 3-kinase/protein kinase B/mammalian target of rapamycin （PI3K/Akt/mTOR） signaling pathway between SMMC-7721 and SMMC-7721/S cells. Furthermore， SMMC-7721/S cells were treated with 5 μmol·L^-1 berbamine hydrochloride alone or in combination with 2.5， 5， 10 μmol·L^-1 sorafenib， and the cell growth was assessed by the CCK-8 assay. In addition， SMMC-7721 and SMMC-7721/S cells were treated with 5 μmol·L^-1 berbamine hydrochloride alone or in combination with 5 μmol·L^-1 sorafenib， and the cell proliferation was examined by the colony formation assay. The immunofluorescence assays with Microtubule-associated protein 1 light chain 3 （LC3） and LysoTracker as probes were employed to assess the lysosomal acidification in SMMC-7721 cells treated with 5 μmol·L^-1 berbamine hydrochloride or 0.1 μmol·L^-1 autophagy inhibitor bafilomycin A1 （Baf）. Further， the expression of proteins involved in autophagy and PI3K/Akt/mTOR signaling pathway was determined by Western blot and compared between groups. ResultSorafenib showed the IC₅₀ of 9.56 mol·L^-1 （P<0.01） and 7.99 mol·L^-1 for SMMC-7721/S and SMMC-7721 cells， respectively， at 24 h. The resistance index （RI） of SMMC-7721/S for sorafenib was 1.20 （P<0.01）， which indicated mild resistance. Compared with SMMC-7721 cells， SMMC-7721/S cells exhibited up-regulated expression of p-mTOR， p-Akt， and LC3Ⅱ， down-regulated expression of p62 protein （P<0.01）， and unchanged Akt protein level. CCK-8 and colony formation assays demonstrated that the combination of berbamine hydrochloride and sorafenib exhibited a synergistic effect （Q>1.15）， with berbamine hydrochloride partially reversing the resistance of liver cancer cells to sorafenib. The immunofluorescence detection of LC3 revealed that berbamine hydrochloride and Baf significantly increased LC3 in SMMC-7721 cells. The detection with LysoTracker as the probe showed that berbamine hydrochloride inhibited the acidity of lysosomes in SMMC-7721 cells （P<0.01）， indicating the suppression of autophagy. Berbamine hydrochloride further enhanced the downregulation of p-mTOR and p-Akt protein levels and did not change the Akt protein level in SMMC-7721 cells exposed to sorafenib. Berbamine hydrochloride inhibited the increase in p-mTOR expression， down-regulated the p-Akt protein level， and did not change the total Akt protein level in the SMMC-7721/S cells exposed to sorafenib. ConclusionBerbamine hydrochloride can ameliorate the resistance of liver cancer cells to sorafenib by inhibiting cellular autophagy and the PI3K/Akt/mTOR signaling pathway.

With the increasing demand for beauty,the treatment of gingival recession has become a common request among patients.Clinically,gingival recession is mainly treated by surgery.The common surgical methods include free gingival grafting,pedicled flap technology and double flap technology(subepithelial connective tissue transplantation combined with coronally advanced flaps).If patients with indications are selected,satisfactory surgical results will be ob-tained.However,there are still some shortcomings in the above mentioned methods,such as the root coverage effect not being satisfactory.In recent years,researchers have put forward some improved schemes to minimize the shortcomings of the above methods to treat different degrees of gingival recession.A gingival unit graft containing gingival papilla and free gingiva can improve the blood supply of the recipient area and improve the effect of root coverage.It can obtain bet-ter root coverage for slight retraction,widening of the angular gingiva and deepening of the vestibular sulcus,but there may be issues with inconsistent color and shape of the gingiva after surgery,as well as poor aesthetic effects.Modified coronally advanced flaps,flaps prepared by the technique of half-thickness,full-thickness and half-thickness,and modi-fied coronally advanced envelope flap technology are designed with the most serious retraction teeth as the center in the case of multiple gingival retractions,both of which can improve the effect of root covering.Tunnel technology and modi-fied tunnel technology,without severing the gingival papilla and tunneling the gingival flap to accommodate the graft,can effectively reduce tissue damage and promote wound healing.This paper reviews the literature and summarizes the outcome of the modified surgery techniques in the treatment of gingival recession.These treatment options for gingival recession are proposed with the aim of improving clinical work,and some suggestions for the treatment of gingival reces-sion to achieve a stable root coverage effect are put forward.In the future,the development direction of mucogingival surgery is to reduce trauma and have a stable curative effect.

Objective To observe the efficacy and safety of repetitive transcranial magnetic stimulation(rTMS)on refractory tinnitus and the differences of resting-state functional magnetic resonance imaging(rs-fMRI)imaging between before and after treatment,and to explore the possible central mechanism of rTMS regulation of tinnitus.Methods Thirty-seven patients with refractory tinnitus admitted in Affiliated Hospital of North Sichuan Medical College from September 2022 to February 2023 were selected and were divided into experimental group(n=20)and control group(n=1 7).The experimental group was given true rTMS treatment,and the control group was given sham stimulation with the same parameters.Tinnitus handicap inventory(THI)score,tinnitus loudness visual analogue scale(VAS)score and rs-fMRI scan were performed before and after treatment.Regional homogeneity(ReHo)was calculated after scanning,and the different brain regions were selected as the area of interest(ROI)and the whole brain functional connection(FC)was performed.Results There were no significant differences in age,gender,education level,tinnitus side,course of disease,hearing level,self-rating depression scale,self-rating anxiety scale the experimental group and control group.There were no significant differences in THI and VAS scores between the two groups before treatment;the THI and VAS scores in the experimental group decreased after 2 weeks of rTMS treatment(P＜0.001),while there was no significant difference in the two scores in the control group before and after treatment.Only 3 patients in the experimental group experienced left facial muscle tremor or transient mild scalp pain during treatment,without other serious side effects.The ReHo of the left cerebellar area 9 increased in the experimental group after rTMS(P＜0.005);the ReHo values in the right inferior temporal gyrus,left posterior central gyrus and left anterior central gyrus increased in the control group after intervention(P＜0.005).The FCs between the right inferior temporal gyrus and the left orbital inferior frontal gyrus,the left anterior cingulate gyrus increased in the experimental group(P＜0.005),and FC between the right inferior temporal gyrus and the left superior marginal gyrus decreased(P＜0.005).The FCs between the right cuneus and the left fusiform gyrus,right superior occipital gyrus decreased in the experimental group after rTMS(P＜0.005).The FC between the right cuneus and the left fusiform gyrus increased in the control group after intervention(P＜0.005),while other FCs remained unchanged.Conclusions rTMS has a certain therapeutic effect on refractory tinnitus with higher safety;regulation of auditory brain network and related non-auditory brain network may be one of the central mechanisms of rTMS treating refractory tinnitus.