ObjectiveTo establish an ultra-performance liquid fingerprint and multi-components determination method for Naomaili granules. To evaluate the quality of different batches by chemometrics， and the anti-neuroinflammatory effects of water extract and main components of Naomaili granules were tested in vitro. MethodsThe similarity and common peaks of 27 batches of Naomaili granules were evaluated by using Ultra performance liquid chromatography （UPLC） fingerprint detection. Ultra-performance liquid chromatography-tandem mass spectrometry （UPLC-MS/MS） technology was used to determine the content of the index components in Naomaili granules and to evaluate the quality of different batches of Naomaili granules by chemometrics. LPS-induced BV-2 cell inflammation model was used to investigate the anti-neuroinflammatory effects of the water extract and main components of Naomaili granules. ResultsThe similarity of fingerprints of 27 batches of samples was > 0.90. A total of 32 common peaks were calibrated， and 23 of them were identified and assigned. In 27 batches of Naomaili granules， the mass fractions of 14 components that were stachydrine hydrochloride， leonurine hydrochloride， calycosin-7-O-glucoside， calycosin，tanshinoneⅠ， cryptotanshinone， tanshinoneⅡ_A， ginsenoside Rb₁， notoginsenoside R₁， ginsenoside Rg₁， paeoniflorin， albiflorin， lactiflorin， and salvianolic acid B were found to be 2.902-3.498， 0.233-0.343， 0.111-0.301， 0.07-0.152， 0.136-0.228， 0.195-0.390， 0.324-0.482， 1.056-1.435， 0.271-0.397， 1.318-1.649， 3.038-4.059， 2.263-3.455， 0.152-0.232， 2.931-3.991 mg∙g^-1， respectively. Multivariate statistical analysis showed that paeoniflorin， ginsenoside Rg₁， ginsenoside Rb₁ and staphylline hydrochloride were quality difference markers to control the stability of the preparation. The results of bioactive experiment showed that the water extract of Naomaili granules and the eight main components with high content in the prescription had a dose-dependent inhibitory effect on the release of NO in the cell supernatant. Among them， salvianolic acid B and ginsenoside Rb₁ had strong anti-inflammatory activity， with IC₅₀ values of （36.11±0.15） mg∙L^-1 and （27.24±0.54） mg∙L^-1， respectively. ConclusionThe quality evaluation method of Naomaili granules established in this study was accurate and reproducible. Four quality difference markers were screened out， and eight key pharmacodynamic substances of Naomaili granules against neuroinflammation were screened out by in vitro cell experiments.

Objective: To fabricate a hydrogel loaded with inositol hexaphosphate-zinc and preliminarily evaluate its performance in self-mineralization and osteoinduction, thereby providing a theoretical basis for the development of bone regeneration materials. Methods: The hydrogel framework (designated DF0) was formed by copolymerizing methacryloyloxyethyltrimethylammonium chloride and four-armed poly(ethylene glycol) acrylate, followed by sequentially loading inositol hexaphosphate anions via electrostatic interaction and zinc ions via chelation. The hydrogel loaded only with inositol hexaphosphate anions was named DF1, while the co-loaded hydrogel was named DF2. The self-mineralization efficacy of the DF0 , DF1 and DF2 hydrogels was characterized using scanning electron microscopy, transmission electron microscopy (TEM), energy dispersive spectroscopy (EDS), and selected area electron diffraction (SAED). The biocompatibility was assessed via live/dead cell staining and a CCK-8 assay. The osteoinductive capacity of the DF0 , DF1 and DF2 hydrogels on MC3T3-E1 cells was assessed via alkaline phosphatase (ALP) and Alizarin Red S (ARS) staining. In the aforementioned cell experiments, cells cultured in standard medium served as the control group Results: The DF0, DF1, and DF2 hydrogels were successfully synthesized. Notably, DF1 and DF2 exhibited distinct self-mineralization within 6 days. Results from TEM, EDS, and SAED confirmed that the mineralization products were amorphous calcium phosphate in group DF1, and amorphous calciumzinc phosphate in group DF2. Biocompatibility tests revealed that none of the hydrogels (DF0, DF1, and DF2) adversely affected cell viability or proliferation. In osteogenic induction experiments, both ALP and ARS staining were intensified in the DF1 and DF2 groups, with the most profound staining observed in the DF2 group. Conclusion The developed inositol hexaphosphate-zinc hydrogel (DF2) demonstrates the dual capacity to generate calcium-phosphate compounds through self-mineralization while exhibiting excellent osteoinductive properties. This biocompatible, dual-promoting osteogenic hydrogel presents a novel strategy for bone regeneration.

BACKGROUND:Neurotrophins represent a novel therapeutic approach for spinal cord injury,showing promising clinical applicability.Autophagy modulation is one of the mechanisms by which neurotrophins exert their effects,yet the specific signaling pathways involved remain unclear. OBJECTIVE:To explore how neurotrophin-3(NT-3)modulates autophagy in oligodendrocytes via switching between P75NTR and TrkC receptors and promotes neurological function recovery after spinal cord injury,aiming to further clarify the specific molecular mechanisms involved. METHODS:Twenty-four Sprague-Dawley rats were randomly divided into three groups:sham operation,spinal cord injury,and NT-3 groups.The therapeutic effect of NT-3 on spinal cord injury in rats was evaluated using the Basso,Beattie,and Bresnahan locomotor rating scale.The expression levels of NT-3,Olig1,myelin basic protein,and the autophagy marker LC3B in rat spinal cord tissue were detected by western blot.In a cellular experiment,oligodendrocytes were cultured in vitro and divided into six groups:oxygen-glucose deprivation(OGD),OGD+NT-3,OGD+NT-3+P75NTR plasmid,OGD+NT-3+TrkC plasmid,OGD+3-methyladenine(an autophagy inhibitor),and OGD+rapamycin(an autophagy activator).Oligodendrocyte morphology was observed under a light microscope,cell apoptosis was assessed by TUNEL staining,and the expression of TrkC receptor,P75NTR,LC3B,and the phosphorylation status of the PI3K/AKT/mTOR and AMPK/mTOR signaling pathways were evaluated by western blot. RESULTS AND CONCLUSION:Animal experiments demonstrated that compared with the sham operation group,NT-3 expression significantly increased after spinal cord injury(P<0.05);exogenous NT-3 treatment accelerated neurological function recovery in rats post spinal cord injury(P<0.05)and increased the expression of Olig1 and myelin basic proteins(P<0.05).Cellular experiments revealed that 3 hours marked the early to middle/late phase transition.Compared with the OGD group,oligodendrocytes in the OGD+NT-3 group could maintain their morphology for a longer period of time,TrkC receptor expression was lower in the early phase and significantly upregulated in the middle/late phase(P<0.05),whereas P75NTR protein expression was upregulated in the early phase and downregulated in the middle/late phase(P<0.05),and autophagy levels showed an initial increase followed by a decrease(P<0.05).By comparing the morphology and TUNEL staining results of cells in the OGD+NT-3,OGD+rapamycin,and OGD+3-methyladenine groups,we found that either promoting or inhibiting autophagy alone had adverse effects on oligodendrocyte survival,whereas modulating autophagy in a manner similar to NT-3 could maximally maintain cell survival.NT-3 could promote autophagy in the early phase via the P75NTR/AMPK/mTOR signaling pathway and inhibit autophagy in the later phase through the TrkC/PI3K/AKT/mTOR signaling pathway.Based on these findings,it is concluded that NT-3 can bidirectionally regulate autophagy in oligodendrocytes through the switching of P75NTR/TrkC receptors,thereby maintaining cell survival and facilitating the recovery of neurological functions in rats after spinal cord injury.

Purpose: Erectile dysfunction (ED) is a common male sexual dysfunction. Gut microbiota plays an important role in various diseases. To investigate the effects and mechanisms of intestinal flora dysregulation induced by high-fat diet (HFD) on erectile function. Materials and Methods: Male Sprague–Dawley rats aged 8 weeks were randomly divided into the normal diet (ND) and HFD groups. After 24 weeks, a measurement of erectile function was performed. We performed 16S rRNA sequencing of stool samples. Then, we established fecal microbiota transplantation (FMT) rat models by transplanting fecal microbiota from rats of ND group and HFD group to two new groups of rats respectively. After 24 weeks, erectile function of the rats was evaluated and 16S rRNA sequencing was performed, and serum samples were collected for the untargeted metabolomics detection. Results: The erectile function of rats and the species diversity of intestinal microbiota in the HFD group was significantly lower, and the characteristics of the intestinal microbiota community structure were also significantly different between the two groups. The erectile function of rats in the HFD-FMT group was significantly lower than that of rats in the ND-FMT group. The characteristics of the intestinal microbiota community structure were significantly different. In the HFD-FMT group, 27 metabolites were significantly different and they were mainly involved in the several inflammation-related pathways. Conclusions Intestinal microbiota disorders induced by HFD can damage the intestinal barrier of rats, change the serum metabolic profile, induce low-grade inflammation and apoptosis in the corpus cavernosum of the penis, and lead to ED.

Background: Kelch-like ECH-associated protein 1 (KEAP1)–nuclear factor erythroid- 2-related factor 2 (NRF2) pathway is a major regulator protecting cells from oxidative and metabolic stress. Studies have revealed that this pathway is involved in mediating resistance to cytotoxic chemotherapy and immunotherapy; however, its implications in oncogene-addicted tumors are largely unknown. This study aimed to elucidate whether this pathway could be a potential therapeutic target for epidermal growth factor receptor (EGFR)-mutant non-small cell lung cancer. Methods: We measured the baseline expression of NRF2 using EGFR-mutant parental cells and acquired gefitinib resistant cells. We investigated whether NRF2 inhibition affected cell death in vitro and tumor growth in vivo using a xenograft mouse model, and compared the transcriptional changes before and after NRF2 inhibition. Results: Baseline NRF2 expression was enhanced in PC9 and PC9 with gefitinib resistance (PC9/GR) cells than in other cell lines, with a more prominent expression in PC9/ GR. The NRF2 inhibitor induced NRF2 downregulation and cell death in a dose-dependent manner. Cotreatment with an NRF2 inhibitor enhanced osimertinib-induced cell death in vitro, and potentiated tumor growth inhibition in a PC9/GR xenograft model. Finally, RNA sequencing revealed that NRF2 inhibition resulted in the altered expression of multiple genes involved in various signaling pathways. Conclusion We identified that NRF2 inhibition enhanced cell death and inhibited tumor growth in tyrosine kinase inhibitor (TKI)-resistant lung cancer with EGFR-mutation. Thus, NRF2 modulation may be a novel therapeutic strategy to overcome the resistance to EGFR-TKIs.

Purpose: The aim of this study was to investigate the feasibility of an intelligent handheld ultrasound (US) device for assisting non-expert general practitioners (GPs) in detecting carotid plaques (CPs) in community populations. Methods: This prospective parallel controlled trial recruited 111 consecutive community residents. All of them underwent examinations by non-expert GPs and specialist doctors using handheld US devices (setting A, setting B, and setting C). The results of setting C with specialist doctors were considered the gold standard. Carotid intima-media thickness (CIMT) and the features of CPs were measured and recorded. The diagnostic performance of GPs in distinguishing CPs was evaluated using a receiver operating characteristic curve. Inter-observer agreement was compared using the intragroup correlation coefficient (ICC). Questionnaires were completed to evaluate clinical benefits. Results: Among the 111 community residents, 80, 96, and 112 CPs were detected in settings A, B, and C, respectively. Setting B exhibited better diagnostic performance than setting A for detecting CPs (area under the curve, 0.856 vs. 0.749; P<0.01). Setting B had better consistency with setting C than setting A in CIMT measurement and the assessment of CPs (ICC, 0.731 to 0.923). Moreover, measurements in setting B required less time than the other two settings (44.59 seconds vs. 108.87 seconds vs. 126.13 seconds, both P<0.01). Conclusion Using an intelligent handheld US device, GPs can perform CP screening and achieve a diagnostic capability comparable to that of specialist doctors.

Purpose: Cognitive behavioral therapy (CBT) has long been recognized as an effective treatment for depression and suicidality.Virtual reality (VR) technology is widely used for cognitive training for conditions such as anxiety disorder and post-traumatic stress disorder, but little research has considered VR-based CBT for depressive symptoms and suicidality. We tested the effectiveness and safety of a VR-based CBT program for depressive disorders. Materials and Methods: We recruited 57 participants from May 2022 through February 2023 using online advertisements. This multi-center, assessor-blinded, randomized, controlled exploratory trial used two groups: VR treatment group and treat as usual (TAU) group. VR treatment group received a VR mental health training/education program. TAU group received standard pharmacotherapy. Assessments were conducted at baseline, immediately after the 6-week treatment period, and 4 weeks after the end of the treatment period in each group. Results: Depression scores decreased significantly over time in both VR treatment and TAU groups, with no differences between the two groups. The suicidality score decreased significantly only in VR group. No group differences were found in the remission or response rate for depression, perceived stress, or clinical severity. No adverse events or motion sickness occurred during the VR treatment program. Conclusion VR CBT treatment for major depressive disorder has the potential to be equivalent to the gold-standard pharmacotherapy in reducing depressive symptoms, suicidality, and related clinical symptoms, with no difference in improvement found in this study. Thus, VR-based CBT might be an effective alternative to pharmacotherapy for depressive disorders.

Purpose: This study aimed to evaluate the prognosis of the not evaluated (NE) group by comparing it with the lost to follow-up (LTFU) group among patients with multidrug/rifampin-resistant tuberculosis (MDR/RR-TB). Materials and Methods: This was a retrospective longitudinal follow-up study using an integrated database constructed by data linkage of the three national databases. This database included 7226 cases of MDR/RR-TB notified between 2011 and 2017 in South Korea. Results: Among the 7226 MDR/RR-TB cases, 730 (10.1%) were classified as LTFU group, and 353 (4.9%) as NE group. When comparing NE group with LTFU group, there were no significant differences in the all-cause mortality rate (18.1% vs. 13.8%, p=0.065), median time to death [404 days (interquartile range, IQR 46–850) vs. 443 days (IQR 185–1157), p=0.140], and retreatment rate (26.9% vs.22.2%, p=0.090). After adjusting for potential confounders, the adjusted hazard ratio (aHR) for all-cause mortality (aHR 1.11; 95% confidence interval 0.80-1.53; p=0.531) in NE group was not significantly different than that in LTFU group. Among retreated cases, NE group had a higher treatment success rate (57.9% vs 43.8%, p=0.029) and a lower LTFU rate (11.6% vs 38.3%, p<0.001) compared to LTFU group. Conclusion NE group had an unfavorable outcome comparable to LTFU group, suggesting undetected cases of LTFU or deaths during the referral process. Establishing an efficient patient referral system would contribute to reducing the incidence of NE cases.

Background: s/Aims: Hepatocellular carcinoma (HCC) is a malignant cancer with an increasing incidence worldwide. Although numerous efforts have been made to identify effective therapies for HCC, current strategies have limitations. We present a new approach for targeting L-arginine and argininosuccinate synthetase 1 (ASS1). Methods: ASS1 expression in HCC cell lines and primary hepatocytes was detected using polymerase chain reaction and western blotting. Proliferation, migration, signaling pathways, and nitric oxide production in HCC cell lines were measured using MTS, colony formation, wound healing, Western blot, and Griess assays. Results: ASS1 expression varied among the HCC cell lines, and cisplatin cytotoxicity was ASS1-dependent. L-arginine alone induced apoptosis in HCC cell lines, regardless of ASS1 expression; however, its effect was enhanced in ASS1-expressing HCC cell lines. Cisplatin cytotoxicity also increased, suggesting that L-arginine acts as a sensitizer to cisplatin in HCC cell lines. ASS1 and L-arginine produced nitric oxide and inhibited key proliferation- and survival-related signaling pathways such as PI3K/Akt and MAPK. Additionally, ASS1 and L-arginine reduced the expression of PKM1 and PKM2 in the glycolysis pathway. Conclusions Our study revealed that ASS1 and L-arginine exhibited anticancer effects in HCC and sensitized cisplatin-resistant HCC cells to chemotherapy. The combination of ASS1 and L-arginine significantly enhanced the anticancer effects, even in HCC cell lines with low or absent ASS1 expression. These findings highlight the critical roles of arginine and ASS1 in HCC and suggest that increasing arginine availability could be a promising therapeutic strategy.

ObjectiveTo explore the effects of different dosage forms of Kaixinsan （KXS） on the morphology and function of mitochondria in rat models of Alzheimer's disease （AD） and potential mechanisms of action. MethodsMale SD rats were randomly assigned to a sham group， model group， treatment groups receiving KXS decoction， powders， and granules （3.08 g·kg^-1）， as well as donepezil group （0.51×10^-3 g·kg^-1）， with 10 rats in each group. AD model was created using intracerebroventricular injection of streptozocin （STZ）. After 30 days of administration， behavioral assessments were conducted， and mitochondrial morphology was observed using transmission electron microscopy. Mitochondrial respiratory chain complex content was measured via enzyme-linked immunosorbent assay （ELISA）. Changes in mitochondrial membrane potential were measured via JC-1 staining， and superoxide dismutase （SOD） activity and reactive oxygen species （ROS） levels were measured via biochemical assays. The mRNA expression of adenosine 5'-monophosphate-activated protein kinase （AMPK）， peroxisome proliferator-activated receptor gamma coactivator-1α （PGC-1α）， and silent information regulator 3 （SIRT3） was detected by real-time fluorescent quantitative polymerase chain reaction （Real-time PCR）， and Western blot was used to examine the protein expression levels of optic atrophy protein1 （OPA1）， mitochondrial fission protein 1 （FIS1）， AMPK， p-AMPK， PGC-1α， and SIRT3. ResultsCompared with the sham group， rats in the model group had significantly lower recognition index， spontaneous alternation rate， escape latency， number of platform crossings， time spent in the target quadrant， and percentage of distance traveled in the target quadrant distance （P<0.05， P<0.01）. Significant mitochondrial damage was observed in the hippocampal tissue， with a marked decrease in mitochondrial respiratory chain complex content （P<0.01） and reduced mitochondrial membrane potential （P<0.05）. Additionally， the SOD activity was reduced， while ROS levels were elevated （P<0.01）. The mRNA expression of PGC-1α and SIRT3 was significantly downregulated （P<0.01）， along with decreased protein expression levels of OPA1， p-AMPK/AMPK， PGC-1α， and SIRT3， whereas FIS1 protein expression was significantly upregulated （P<0.05， P<0.01）. Compared with the model group， rats in KXS-treated groups （various dosage forms） showed significant improvement in behavioral indexes （P<0.05， P<0.01）， reduced hippocampal mitochondrial damage， and more organized mitochondrial cristae. Mitochondrial respiratory chain complex content was significantly increased （P<0.05， P<0.01）， and mitochondrial membrane potentials were elevated （P<0.05）. SOD activity was elevated， and ROS levels were significantly reduced （P<0.05， P<0.01）. Furthermore， the mRNA expression of PGC-1α and SIRT3 was upregulated， with increased protein levels of OPA1， p-AMPK/AMPK， PGC-1α， and SIRT3， while FIS1 protein expression levels were significantly reduced （P<0.05， P<0.01）. Across the KXS-treated groups， the granule group showed a higher spontaneous alternation rate than the decoction and powder groups （P<0.05）. ConclusionKXS decoction， powders， and granules can improve the learning and memory ability of rats， with granules being the most effective. The mechanism of action may involve activation of the AMPK/PGC-1α/SIRT3 signaling pathway， improvement of the mitochondrial function， and subsequent amelioration of the brain energy metabolism disorders.