Periodontitis is a common oral disease caused by bacteria coupled with an excessive host immune response. Stem cell therapy can be a promising treatment strategy for periodontitis, but the relevant mechanism is complicated. This study aimed to explore the therapeutic potential of mitochondria from human embryonic stem cell-derived mesenchymal stem cells (hESC-MSCs) for the treatment of periodontitis. The gingival tissues of periodontitis patients are characterized by abnormal mitochondrial structure. Human gingival fibroblasts (HGFs) were exposed to 5 μg/mL lipopolysaccharide (LPS) for 24 h to establish a cell injury model. When treated with hESC-MSCs or mitochondria derived from hESC-MSCs, HGFs showed reduced expression of inflammatory genes, increased adenosine triphosphate (ATP) level, decreased reactive oxygen species (ROS) production, and enhanced mitochondrial function compared to the control. The average efficiency of isolated mitochondrial transfer by hESC-MSCs was determined to be 8.93%. Besides, a therapy of local mitochondrial injection in mice with LPS-induced periodontitis showed a reduction in inflammatory gene expression, as well as an increase in both the mitochondrial number and the aspect ratio in gingival tissues. In conclusion, our results indicate that mitochondria derived from hESC-MSCs can reduce the inflammatory response and improve mitochondrial function in HGFs, suggesting that the transfer of mitochondria between hESC-MSCs and HGFs serves as a potential mechanism underlying the therapeutic effect of stem cells.
Humans ; Gingiva/cytology* ; Fibroblasts/metabolism* ; Mitochondria/physiology* ; Mesenchymal Stem Cells/cytology* ; Animals ; Periodontitis/therapy* ; Mice ; Reactive Oxygen Species/metabolism* ; Inflammation ; Lipopolysaccharides ; Human Embryonic Stem Cells/cytology* ; Cells, Cultured ; Adenosine Triphosphate/metabolism* ; Male

Despite therapy with potent antiviral agents, chronic hepatitis B (CHB) patients remain at high risk of hepatocellular carcinoma (HCC). While metabolites have been rediscovered as active drivers of biological processes including carcinogenesis, the specific metabolites modulating HCC risk in CHB patients are largely unknown. Here, we demonstrate that baseline plasma from CHB patients who later developed HCC during follow-up exhibits growth-promoting properties in a case-control design nested within a large-scale, prospective cohort. Metabolomics analysis reveals a reduction in long-chain acylcarnitines (LCACs) in the baseline plasma of patients with HCC development. LCACs preferentially inhibit the proliferation of HCC cells in vitro at a physiological concentration and prevent the occurrence of HCC in vivo without hepatorenal toxicity. Uptake and metabolism of circulating LCACs increase the intracellular level of acetyl coenzyme A, which upregulates histone H3 Lys14 acetylation at the promoter region of KLF6 gene and thereby activates KLF6/p21 pathway. Indeed, blocking LCAC metabolism attenuates the difference in KLF6/p21 expression induced by baseline plasma of HCC/non-HCC patients. The deficiency of circulating LCACs represents a driver of HCC in CHB patients with viral control. These insights provide a promising direction for developing therapeutic strategies to reduce HCC risk further in the antiviral era.

Obesity usually exacerbates the immunosuppressive tumor microenvironment (ITME), hindering CD8⁺ T cell infiltration and function, which further represents a significant barrier to the efficacy of immunotherapy. Herein, a multifunctional liposomal system (CR-Lip) for encapsulating celastrol (CEL) was utilized to remodel obesity-related ITME and improve cancer immunotherapy, wherein Ginsenoside Rg3 (Rg3) was detected interspersed in the phospholipid bilayer and its glycosyl exposed on the surface of the liposome. CR-Lip had a relatively uniform size (116.5 nm), facilitating favorable tumor tissue accumulation through the interaction between Rg3 and glucose transporter 1 overexpressed in obese tumor cells. Upon reaching the tumor region, CR-Lip was found to induce the immunogenic cell death (ICD) of HFD tumor cells. Notably, the level of PHD3 in HFD tumor cells was effectively boosted by CR-Lip to effectively block metabolic reprogramming and increase the availability of major free fatty acids fuel sources. In vivo, experiments studies revealed that the easy-obtained nano platform stimulated enhanced the production of various cytokines in tumor tissues, DC maturation, CD8⁺ T-cell infiltration, and synergistic anticancer therapeutic potency with aPD-1 (tumor inhibition rate = 82.1%) towards obesity-related melanoma. Consequently, this study presented an efficacious approach to tumor immunotherapy in obese mice by encompassing tumor eradication, inducing ICD, and reprogramming metabolism. Furthermore, it offered a unique insight into a valuable attempt at the immunotherapy of obesity-associated related tumors.

The interest in covalent drugs has resurged in recent decades, spurring the development of numerous specialized computational docking tools to facilitate covalent ligand design and screening. Herein, we present CarsiDock-Cov, a new paradigm distinguishing itself as the first deep learning (DL)-guided approach for covalent docking. CarsiDock-Cov retains the core components of its non-covalent predecessor, leveraging a DL model pretrained on millions of docking complexes to predict protein-ligand distance matrices, along with a dedicated-designed geometric optimization procedure to convert these distances into refined binding poses. Additionally, it incorporates several key enhancements specifically tailored to optimize the protocol for covalent docking applications. Our approach has been extensively validated on multiple public datasets regarding the docking and screening of covalent ligands, and the results indicate that our approach not only achieves comparably improved applicability compared to its non-covalent predecessor, but also exhibits competitive performance against various state-of-the-art covalent docking tools. Collectively, our approach represents a significant advance in covalent docking methodology, offering an automated and efficient solution that shows considerable promise for accelerating covalent drug discovery and design.

OBJECTIVE: To explore the preventive effect of transcutaneous phrenic nerve stimulation on ventilator-induced diaphragmatic dysfunction (VIDD) in patients requiring invasive mechanical ventilation. METHODS: A randomized controlled trial was conducted. The patients requiring invasive mechanical ventilation admitted to the intensive care unit (ICU) of Jiaxing First Hospital from November 2022 to December 2023 were enrolled. Participants were randomized into the control group and the observation group using a random number table. The control group was given ICU standardized nursing intervention, including turning over and slapping the back, raising the head of the bed, sputum aspiration on demand, aerosol inhalation, oral care, and monitoring of airbag pressure and gastric retention, the observation group was given additional transcutaneous phrenic nerve stimulation intervention on the basis of ICU standardized nursing intervention. The stimulation intensity was set to 10 U, the pulse frequency was set to 40 Hz, and the stimulation frequency was set to 12 times/min. Transcutaneous phrenic nerve stimulation was administered once a day for 30 minutes each time, for a total of 5 days. Diaphragm thickening fraction (DTF) and arterial blood gas parameters on days 1, 3, and 5 of intervention were compared between the two groups. After 5 days of intervention, other parameters including the incidence of VIDD, duration of mechanical ventilation, and length of ICU stay were compared. RESULTS: A total of 120 patients requiring invasive mechanical ventilation were enrolled, with 16 dropouts (dropout rate was 13.33%). Ultimately, 51 patients in the control group and 53 patients in the observation group were analyzed. Baseline characteristics, including gender, age, body mass index (BMI), acute physiology and chronic health evaluation II (APACHE II) score, albumin (Alb), hemoglobin (Hb), and disease type, showed no significant differences between the two groups. DTF in both groups gradually increased over duration of intervention [DTF on days 1, 3, and 5 in the control group was (20.83±2.33)%, (21.92±1.27)%, and (23.93±2.33)%, respectively, and that in the observation group was (20.89±1.96)%, (22.56±1.64)%, and (25.34±2.38)%, respectively], with more significant changes in DTF in the observation group, showing time effects (F_time = 105.975, P < 0.001), intervention effects (F_intervention = 7.378, P = 0.008), and interaction effects (F_interaction = 3.322, P = 0.038). Arterial blood gas parameters did not differ significantly before intervention between the groups, but after 5 days of intervention, arterial partial pressure of oxygen (PaO₂) in the observation group was significantly higher than that in the control group [mmHg (1 mmHg≈0.133 kPa): 100.72±15.75 vs. 93.62±15.54, P < 0.05], and arterial partial pressure of carbon dioxide (PaCO₂) was significantly lower than that in the control group (mmHg: 36.53±3.10 vs. 37.69±2.02, P < 0.05). At 5 days of intervention, the incidence of VIDD in the observation group was significantly lower than that in the control group [15.09% (8/53) vs. 37.25% (19/51), P < 0.05], and both duration of mechanical ventilation and length of ICU stay were significantly shorter than those in the control group [duration of mechanical ventilation (days): 7.93±2.06 vs. 8.77±1.76, length of ICU stay (days): 9.64±2.35 vs. 11.01±2.01, both P < 0.05]. CONCLUSIONS Transcutaneous phrenic nerve stimulation can improve diaphragmatic and respiratory function in patients receiving invasive mechanical ventilation, reduce the incidence of VIDD, and shorten the duration of mechanical ventilation and length of ICU stay.
Humans ; Transcutaneous Electric Nerve Stimulation ; Respiration, Artificial/adverse effects* ; Diaphragm/physiopathology* ; Phrenic Nerve ; Intensive Care Units ; Male ; Female ; Middle Aged

Objective:To explore the clinical efficacy of areola approach endoscopic thyroidectomy (AET) and gasless axillary approach endoscopic thyroidectomy (GAET) in the treatment of papillary thyroid carcinoma (PTC) patients.Methods:A total of 96 PTC patients from the Thyroid Surgery Department of Linyi People’s Hospital from May. 2019 to May. 2022 were selected and randomly divided into 48 patients using a random number table method. The areola group received AET, while the armpit group received GAET. The surgical situation, postoperative recovery, relevant biochemical indicators [white blood cell count (WBC), erythrocyte sedimentation rate (ESR), C-reactive protein (CRP), parathyroid hormone (PTH), blood calcium] before and after surgery, postoperative pain level, discomfort level, neck function, and complications were compared between the two groups.Results:The surgical time and extubation time of the armpit group were (125.71±15.73) minutes and (3.12±0.53) days, respectively, which were shorter than those of the areola group (137.94±20.02) minutes and (3.48±0.46) days. The intraoperative bleeding volume was (14.19±4.16) mL, which was less than that of the areola group (22.65±7.39) mL, and the number of lymph nodes cleaned was 5.06±1.02, which was more than that of the areola group (4.23±1.14) ( P<0.05) ; there was no significant difference in postoperative drainage volume and hospital stay between the two groups ( P>0.05) ; Peripheral blood WBC in the armpit group on the 1st and 3rd day after surgery [ (5.69±0.15) ×10 9/L, (5.52±0.14) ] ×10 9/L, ESR [ (8.21±0.55) mm/h, (7.64±0.60) mm/h], CRP [ (10.06±1.78) ng/L, (8.93±1.33) ng/L] were lower than those in the areola group [ (5.83±0.21) ×10 9/L, (5.70±0.23) ×10 9/L, (8.87±0.74) mm/h, (8.19±0.68) mm/h, (12.45±1.90) ng/L, (10.45±1.50) ng/L] ( P<0.05). There was no significant difference in the levels of the above biochemical indicators 5 days after surgery ( P>0.05). There was no significant difference in peripheral blood PTH and calcium levels between the two groups on the 1st, 3rd, and 5th postoperative days ( P>0.05). The pain level [ (3.25±0.32) scores, (2.53±0.27) scores, (1.82±0.22) scores] and discomfort level [ (6.85±0.71) scores, (5.24±0.66) scores, (3.51±0.57) scores] in the axillary group were lower than those in the areola group [ (3.78±0.40) scores, (2.89±0.34) scores, (2.06±0.26) scores, (7.46±0.84) scores, (6.09±0.73) scores, (4.16±0.60) scores] on the 1st, 3rd, and 5th postoperative days ( P<0.05). The neck flexion, lateral flexion, and extension range of motion in the axillary group on the 3rd day after surgery were (33.16±3.09) °, (27.63±2.57) °, and (30.44 2.73) °, respectively, which were greater than those in the areola group[ (30.08±2.76) °, (25.14±2.30) °, and (27.98±2.54) °], and the swallowing disorder index was (30.16±4.97) points lower than the (34.83±4.13) points in the areola group ( P<0.05). The incidence of complications in the axillary group was 4.17% (2/48), lower than the 16.67% (8/48) in the areola group. Conclusion:GAET treatment for PTC patients can improve the effect of lymph node dissection, reduce the degree of surgical trauma, postoperative pain and discomfort, accelerate early postoperative recovery of neck function, and reduce complications.

OBJECTIVE To provide reference for improving the rational use of antimicrobial drugs in primary township medical institutions. METHODS Based on the county prescription pre-review center， a team led by anti-infective clinical pharmacists constructed the management mode for the use of antimicrobial drugs in county-level medical communities with clinical pharmacists as the main team by finding out the main problems in the use of antimicrobial drugs in primary township medical institutions， providing feedback on the problems， organizing relevant training for the problems， improving the customization rules of the prescription pre-review software， implementing the automatic interception and pharmacist online prescription review and other measures. Data on the use of antimicrobial drugs were collected and compared in the 15 primary township medical institutions between January-June in 2022 （before the implementation of the mode） and January-June in 2023 （after the implementation of the mode）. RESULTS Compared with before the implementation of the mode， the utilization rate of antimicrobial drugs in outpatients of primary township medical institutions decreased from 24.97% before the implementation of the mode to 19.39% after the implementation of the mode； the utilization rate of antimicrobial injection in outpatients decreased from 66.10% to 46.80%； the utilization rate of intravenous drip of antimicrobial drugs in outpatients decreased from 52.33% to 40.35%； the rates of combined use of antimicrobial drugs in outpatients decreased from 12.70% to 8.19%； the reasonable rate of antimicrobial prescribing in outpatients increased from 55.28% to 73.93%. After the implementation of the mode， the proportion of antimicrobial prescriptions for each diagnosis was basically the same as before； the defined daily dose system （DDDs） and proportion of a few antimicrobial drugs changed compared with before according to the anatomical therapeutic chemical classification of drugs， among which DDDs of lincomycin， gentamicin and other drugs declined significantly； DDDs of antimicrobial drugs for each classification was basically the same as before according to AWaRe classification. CONCLUSIONS The management mode of the use of antimicrobial drugs led by anti-infective clinical pharmacists is constructed in the prescription pre-reviewing center of county-level medical communities， which can effectively improve the rational use of antimicrobial drugs in the primary township medical institutions.

Objective: To explore the neuroprotective effects of the Shaoyao Gancao decoction (SGD) against excitatory damage in PC12 cells and the role of the Src-NR2-nNOS pathway mediation by SGD in regulating γ-aminobutyric acid (GABA)-glutamate (Glu) homeostasis. Methods: N-Methyl-d-aspartic acid (NMDA) was used to establish a PC12 cell excitability injury model. To investigate the neuroprotective effect of SGD, a cell counting kit-8 (CCK-8) assay was used to determine PC12 cell viability, Annexin V/Propidium Iodide (Annexin V/PI) double staining was used to determine PC12 cell apoptosis, and Ca2+ concentration was observed using laser confocal microscopy. GABA receptor agonists and antagonists were used to analyze the neuroprotective interactions between γ-aminobutyric acid (GABA) and NMDA receptors. Additionally, molecular biology techniques were used to determine mRNA and protein expression in the Src-NR2-nNOS pathway. We analyzed the correlations between the regulatory sites of GABA and NMDA interactions, excitatory neurotoxicity, and brain damage at the molecular level. Results: NMDA excitotoxic injury manifested as a significant decrease in cell activity, increased apoptosis and caspase-3 protein expression, and a significant increase in intracellular Ca2+ concentration. Administration of SGD, a GABAA receptor agonist (muscimol), or a GABAB receptor agonist (baclofen) decreased intracellular Ca2+ concentrations, attenuated apoptosis, and reversed NMDA-induced upregulation of caspase-3, Src, NMDAR2A, NMDAR2B, and nNOS. Unexpectedly, a GABAA receptor antagonist (bicuculline) and a GABAB receptor antagonist (saclofen) failed to significantly increase excitatory neurotoxicity. Conclusions Taken together, these results not only provide an experimental basis for SGD administration in the clinical treatment of central nervous system injury diseases, but also suggest that the Src-NR2A-nNOS pathway may be a valuable target in excitotoxicity treatment.

Objective:To investigate the role and underlying mechanisms of methyltransferase (Mettl) 3 in the process of angiotensin Ⅱ (Ang Ⅱ)-induced pericyte-to-myofibroblast transdifferentiation and renal fibrosis.Methods:C57BL/6J mice were used, in cell experiments, mouse renal pericytes were isolated and cultured using magnetic bead sorting. These pericytes were then induced to transdifferentiate into myofibroblasts with 1×10 6 mmol/L Ang Ⅱ, which was the Ang Ⅱ group, while pericytes cultured in normal conditions served as the control group. Successful transdifferentiation was verified by immunofluorescence staining, Western blotting, and real-time reverse transcription PCR (RT-qPCR) for α-smooth muscle actin (α-SMA). The levels of m6A modifications and related enzymes (Mettl3, Mettl14), Wilms tumor 1-associated protein (WTAP), fat mass and obesity protein (FTO), ALKBH5, YTHDF1, YTHDF2, YTHDC1, YTHDC2, YTHDC3 were assessed by Dot blot, RT-qPCR and Western blot. Mettl3 expression was inhibited in cells using lentivirus-mediated Mettl3-shRNA transfection, creating sh-Mettl3 and Ang Ⅱ+sh-Mettl3 groups, while lentivirus empty vector transfection served as the negative control (Ang Ⅱ+sh-NC group). The impact of Ang Ⅱ on pericyte transdifferentiation was observed, and the expression of downstream phosphatidylinositol 3-kinase (PI3K)/AKT signaling pathway proteins, including PI3K, AKT, phosphorylated AKT at serine 473 (p-AKT (S473)), and phosphorylated AKT at threonine 308 (p-AKT (T308)), were examined. PI3K gene transcription was inhibited by co-culturing cells with actinomycin D, and the half-life of PI3K mRNA was calculated by measuring residual PI3K mRNA expression over different co-culture time. The reversibility of Mettl3 inhibition on Ang Ⅱ-induced pericyte-to-myofibroblast transdifferentiation was assessed by adding the AKT activator SC79 to the Ang Ⅱ+sh-Mettl3 group. In animal experiments, mice were divided into these groups: sham group (administered 0.9% sterile saline), Ang Ⅱ group (infused with Ang Ⅱ solution), sh-Mettl3 group (injected with Mettl3 shRNA lentivirus solution), Ang Ⅱ+sh-Mettl3 group (infused with Ang Ⅱ solution and injected with Mettl3 shRNA lentivirus solution), and Ang Ⅱ+sh-Mettl3+SC79 group (administered Ang Ⅱ solution and Mettl3 shRNA lentivirus, with an additional injection of SC79). Each group consisted of six subject mice. Blood pressure was measured using the tail-cuff method before and after surgery, and serum creatinine, urea, and urinary albumin levels were determined 4 weeks post-surgery. Kidney tissues were collected at 28 days and stained using hematoxylin-eosin (HE) and Masson′s trichrome to assess the extent of renal fibrosis. Results:Primary renal pericytes were successfully obtained by magnetic bead sorting, and intervened with 1×10 6 mmol/L Ang Ⅱ for 48 hours to induce pericyte-to-myofibroblast transdifferentiation. Dot blot results indicated higher m6A modification levels in the Ang Ⅱ group compared to the control group ( P<0.05). RT-qPCR and Western blot results showed upregulation of Mettl3 mRNA and protein levels in the Ang Ⅱ group compared to the control group (both P<0.05). In the Ang Ⅱ+sh-Mettl3 group, Mettl3 protein expression was lower than that in the Ang Ⅱ group, with reduced expression levels of α-SMA, vimentin, desmin, fibroblast agonist protein (FAPa) and type Ⅰ collagen (all P<0.05). Compared to the control group, PI3K mRNA expression level was elevated in the Ang Ⅱ group, along with increased p-AKT (S473) and p-AKT (T308) expressions. In the Ang Ⅱ+sh-Mettl3 group, PI3K mRNA expression and p-AKT (S473) and p-AKT (T308) levels were decreased (all P<0.05). The half-life of PI3K mRNA was shorter in the Ang Ⅱ+sh-Mettl3 group than that in the Ang Ⅱ+sh-NC group (2.34 h vs. 3.42 h). The ameliorative effect of Mettl3 inhibition on Ang Ⅱ-induced pericyte-to-myofibroblast transdifferentiation was reversible by SC79. Animal experiments showed higher blood pressure, serum creatinine, urea, and 24-hour urinary protein levels, and a larger fibrosis area in the Ang Ⅱ group compared to the sham group (all P<0.05). The fibrosis area was smaller in the Ang Ⅱ+sh-Mettl3 group than that in the Ang Ⅱ group ( P<0.05), but increased again upon addition of SC79. Conclusion:Mettl3-mediated RNA m6A epigenetic regulation is involved in Ang Ⅱ-induced pericyte-to-myofibroblast transdifferentiation and renal fibrosis, potentially by affecting PI3K stability and regulating the PI3K/AKT signaling pathway.

Background and objective Invasive mucinous adenocarcinoma(IMA)was a rare and specific type of lung adenocarcinoma,which was often characterized by fewer lymphatic metastases.Therefore,it was difficult to evaluate the prognosis of these tumors based on the existing tumor-node-metastasis(TNM)staging.So,this study aimed to develop Nomo-grams to predict outcomes of patients with pathologic N0 in resected IMA.Methods According to the inclusion criteria and exclusion criteria,IMA patients with pathologic N0 in The Affiliated Lihuili Hospital of Ningbo University(training cohort,n=78)and Ningbo No.2 Hospital(validation cohort,n=66)were reviewed between July 2012 and May 2017.The prognostic value of the clinicopathological features in the training cohort was analyzed and prognostic prediction models were established,and the performances of models were evaluated.Finally,the validation cohort data was put in for external validation.Results Univariate analysis showed that pneumonic type,larger tumor size,mixed mucinous/non-mucinous component,and higher overall stage were significant influence factors of 5-year progression-free survival(PFS)and overall survival(OS).Multivariate analysis further indicated that type of imaging,tumor size,mucinous component were the independent prognostic factors for poor 5-year PFS and OS.Moreover,the 5-year PFS and OS rates were 62.82％and 75.64％,respectively.In subgroups,the sur-vival analysis also showed that the pneumonic type and mixed mucinous/non-mucinous patients had significantly poorer 5-year PFS and OS compared with solitary type and pure mucinous patients,respectively.The C-index of Nomograms with 5-year PFS and OS were 0.815(95％CI:0.741-0.889)and 0.767(95％CI:0.669-0.865).The calibration curve and decision curve analysis(DCA)of both models showed good predictive performances in both cohorts.Conclusion The Nomograms based on clinicopathological characteristics in a certain extent,can be used as an effective prognostic tool for patients with pathologic N0 after IMA resection.