ObjectiveTo observe the intervention effect of Fufang Kangjiaolv capsules on anxiety-like behaviors in the rat model of chronic restraint stress （CRS） and explore the mechanism underlying the anti-anxiety effect via the microbiota-gut-brain axis. MethodsRats were assigned into blank， model， positive drug （diazepam， 1 mg·kg^-1）， and low-， medium-， and high-dose （0.75， 1.5， 3 g·kg^-1， respectively） Fufang Kangjiaolv capsules groups. After 14 days of administration， the elevated plus maze test， open field test， light and dark box test， and marble burying test were performed. Hematoxylin-eosin staining was employed to observe the pathological changes in the hippocampus and colon of rats， and Nissl staining was conducted to observe the damage of hippocampal neurons. The gut microbiota was analyzed by 16S rRNA gene sequencing. Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） was employed to determine the mRNA levels of zonula occludens-1 （ZO-1） and occludin in the colon of rats. The levels of tumor necrosis factor-α （TNF-α）， interleukin-6 （IL-6）， and interleukin-1β （IL-1β） in the colon， serum， and hippocampus were determined by enzyme-linked immunosorbent assay. Western blot was employed to determine the protein levels of ZO-1， occludin， nuclear factor-κB p65 （NF-κB p65） in the colon tissue and NF-κB p65 and brain-derived neurotrophic factor （BDNF） in the hippocampal tissue. ResultsCompared with the blank group， the model group showed reductions in the time and frequency ratio of rats entering the elevated plus maze， the time and frequency of rats entering the central area of the open field， the time of entering the open box， the times of passing through the light and dark box， and the number of unburied beads （P<0.05， P<0.01）. Compared with the model group， Fufang Kangjiaolv capsules ameliorated the anxiety of the model rats to varying degrees， and the high-dose group had the best effect， with increases in the proportions of time and frequency of rats entering the open arm in the elevated plus maze （P<0.05）， the number of rats entering the central area in the open field （P<0.05）， the time of entering the open box， the times of passing through the light and dark boxes， and the number of unburied beads （P<0.01）. Moreover， the high-dose group showed alleviated pathological damage of hippocampal neurons and colon. The results of 16S rRNA gene sequencing showed that the model group had increased relative abundance of Firmicutes， Deferribacterota， Romboutsia， and Phascolarctobacterium， while it had decreased relative abundance of Bavcteroidota and Lactobacillus. The drug administration groups showed increased relative abundance of Bavcteroidota， Bacteroides， norank f norank o Clostridia UCG-014， and Blautia and decreased relative abundance of Firmicutes and Deferribacterota. Compared with the blank group， the model group showed down-regulated protein and mRNA levels of ZO-1 and occludin in the colon （P<0.01）， elevated levels of TNF-α， IL-6， and IL-β in the colon， serum， and hippocampus （P<0.01）， up-regulated protein level of NF-κB p65 in the colon and hippocampus （P<0.01）， and down-regulated protein level of BDNF in the hippocampus （P<0.05）. Compared with the model group， high-dose Fufang Kangjiaolv capsules up-regulated the mRNA levels of ZO-1 and occludin in the colon （P<0.01）， lowered the levels of TNF-α， IL-6， and IL-β in the colon， serum， and hippocampus （P<0.01）， up-regulated the protein levels of ZO-1 （P<0.01） and occludin （P<0.05） in the colon， down-regulated the protein level of NF-κB p65 in the colon and hippocampus （P<0.05）， and up-regulated the protein level of BDNF in the hippocampus. ConclusionFufang Kangjiaolv capsules can reduce the anxiety-like behaviors in the rat model of CRS by regulating the gut microbiota disturbance， up-regulating the expression of tight junction proteins in the colon， repairing intestinal mucosal mechanical barrier， and down-regulating NF-κB/BDNF signaling pathway， thereby reducing peripheral and central inflammation. This study proves the hypothesis that Fufang Kangjiaolv capsules play an anti-anxiety role via the microbiota-gut-brain axis， providing a new idea for further research.

The organoid co-culture model, as a novel tool for recreating a three-dimensional microenvironment to study cell-cell interactions, has demonstrated significant application potential in biomedical research in recent years. By simulating the in vivo tissue microenvironment, this model provides a more precise experimental platform for investigating complex cellular interactions, particularly in areas such as tumor immune evasion mechanisms, drug sensitivity testing, and the pathological characterization of neurodegenerative diseases, where it has demonstrated significant value. However, the organoid co-culture model still faces several challenges in terms of standardized procedures, large-scale cultivation, ethical guidelines, and future development. In particular, in the field of laboratory animal science, how to effectively combine organoids with traditional animal models, and how to select the most appropriate model for different research needs while exploring its potential for replacement, remain pressing issues. In the context of ethical approval and the replacement of animal experiments, the organoid co-culture model offers an experimental approach that better aligns with the "3R" principle (Replacement, Reduction, Refinement), potentially becoming an important tool for replacing traditional animal models. To this end, this paper reviews the latest advances and key challenges in this field, providing a detailed description of the construction methods for organoid co-culture models and discussing their applications in disease mechanism research and drug screening. The paper also systematically compares the organoid co-culture models with traditional animal models, exploring the criteria for selecting the appropriate model for specific applications. Furthermore, this paper discusses the potential value of organoid co-culture models as alternatives to animal experiments and anticipates future development trends of this technology. Through these discussions, the paper aims to promote the innovation and development of organoid co-culture technology and provide new perspectives and scientific evidence for future research.

The organoid co-culture model, as a novel tool for recreating a three-dimensional microenvironment to study cell-cell interactions, has demonstrated significant application potential in biomedical research in recent years. By simulating the in vivo tissue microenvironment, this model provides a more precise experimental platform for investigating complex cellular interactions, particularly in areas such as tumor immune evasion mechanisms, drug sensitivity testing, and the pathological characterization of neurodegenerative diseases, where it has demonstrated significant value. However, the organoid co-culture model still faces several challenges in terms of standardized procedures, large-scale cultivation, ethical guidelines, and future development. In particular, in the field of laboratory animal science, how to effectively combine organoids with traditional animal models, and how to select the most appropriate model for different research needs while exploring its potential for replacement, remain pressing issues. In the context of ethical approval and the replacement of animal experiments, the organoid co-culture model offers an experimental approach that better aligns with the "3R" principle (Replacement, Reduction, Refinement), potentially becoming an important tool for replacing traditional animal models. To this end, this paper reviews the latest advances and key challenges in this field, providing a detailed description of the construction methods for organoid co-culture models and discussing their applications in disease mechanism research and drug screening. The paper also systematically compares the organoid co-culture models with traditional animal models, exploring the criteria for selecting the appropriate model for specific applications. Furthermore, this paper discusses the potential value of organoid co-culture models as alternatives to animal experiments and anticipates future development trends of this technology. Through these discussions, the paper aims to promote the innovation and development of organoid co-culture technology and provide new perspectives and scientific evidence for future research.

OBJECTIVE: To compare the clinical efficacy of acupuncture with yin-yang regulation method versus local acupuncture in treating chronic low back pain (CLBP) in elderly patients with lumbar disc herniation (LDH), and to evaluate the changes in the multifidus muscle before and after treatment using musculoskeletal ultrasound. METHODS: A total of 128 elderly patients with CLBP due to LDH were randomly assigned to an observation group (64 cases, 2 cases dropped out) and a control group (64 cases, 2 cases dropped out). The control group received local acupuncture at bilateral L₃-L₅ Jiaji points (EX-B2), Shenshu (BL23), Dachangshu (BL25), Weizhong (BL40), Yaoyangguan (GV3), and ashi points. The observation group received acupuncture with yin-yang regulation method, which included an abdominal protocol with Baihui (GV20), Zhongwan (CV12), Qihai (CV6), Guanyuan (CV4), bilateral Tianshu (ST25), and Dahe (KI12), etc., and a lumbar protocol with Baihui (GV20), Dazhui (GV14), Jizhong (GV6), Yaoyangguan (GV3), and ashi points, etc., alternated bilaterally. Both groups were treated once every other day, three times per week, for a total of 12 sessions. The visual analogue scale (VAS) score, Oswestry disability index (ODI) score, and the indexs of musculoskeletal ultrasound multifidus muscle (resting and functional thickness and Young's modulus values) were observed before and after treatment, and the clinical efficacy was evaluated in the two groups. RESULTS: After 1 and 4 weeks of treatment, both groups showed lower VAS scores compared to baseline (P<0.05), the VAS scores in the observation group were lower than those in the control group (P<0.001). ODI scores in both groups were decreased after 1 and 4 weeks of treatment compared to baseline (P<0.05), with a further reduction at 4 weeks of treatment compared to 1 week of treatment (P<0.05); the observation group showed lower ODI score than the control group after 1 week of treatment (P<0.001). After treatment, both groups demonstrated increased resting and functional multifidus muscle thickness bilaterally compared to baseline (P<0.01), with an increased right-side thickness change rate (P<0.01), though no significant difference was observed between groups (P>0.05). Compared to baseline, after treatment, the observation group exhibited decreased Young's modulus values for bilateral resting and functional multifidus muscle (P<0.01), while the control group showed reductions only in bilateral resting and right-side functional Young's modulus values (P<0.01). After treatment, the bilateral functional Young's modulus values in the observation group were lower than that in the control group (P<0.05), and the bilateral resting and functional changes in Young's modulus values were greater in the observation group than those in the control group (P<0.01). The overall effective rate was 93.5% (58/62) in the observation group, which was higher than 79.0% (49/62) in the control group (P<0.05). CONCLUSION Acupuncture with yin-yang regulation method effectively alleviates pain, improves functional disability, increases multifidus muscle thickness, and reduces Young's modulus values in elderly patients with CLBP due to LDH, which has superior therapeutic effect compared to local acupuncture.
Humans ; Low Back Pain/physiopathology* ; Male ; Acupuncture Therapy ; Female ; Aged ; Intervertebral Disc Displacement/physiopathology* ; Middle Aged ; Yin-Yang ; Lumbar Vertebrae ; Acupuncture Points ; Treatment Outcome

Nucleotide-binding oligomerization domain (NOD)-like receptor protein 3 (NLRP3) is a cytosolic pattern recognition receptor that recognizes multiple pathogen-associated molecular patterns and damage-associated molecular patterns. It is a cytoplasmic immune factor that responds to cellular stress signals, and it is usually activated after infection or inflammation, forming an NLRP3 inflammasome to protect the body. Aberrant NLRP3 inflammasome activation is reportedly associated with some inflammatory diseases and metabolic diseases. Recently, there have been mounting indications that NLRP3 inflammasomes play an important role in liver injuries caused by a variety of diseases, specifically hepatic ischemia/reperfusion injury, hepatitis, and liver failure. Herein, we summarize new research pertaining to NLRP3 inflammasomes in hepatic injury, hepatitis, and liver failure. The review addresses the potential mechanisms of action of the NLRP3 inflammasome, and its regulation in these liver diseases.
Humans ; NLR Family, Pyrin Domain-Containing 3 Protein/metabolism* ; Inflammasomes/physiology* ; Animals ; Liver Diseases/metabolism* ; Liver/metabolism* ; Reperfusion Injury/metabolism*

Soybean meal (SBM) prepared by soybean crushing is the most popular protein source in the poultry and livestock industries (Cai et al., 2015) due to its economic manufacture, high protein content, and good nutritional value. Despite these benefits, SBM contains various antigen proteins such as glycinin and β-conglycinin, which account for approximately 70% of the total proteins of the SBM and reduce digestibility and damage intestinal function (Peng et al., 2018). Treating SBM with proteases (neutrase, alcalase, and trypsin) or fermentation can eliminate these antigen proteins (Contesini et al., 2018). Because of its safety and rapid growth cycle, Bacillus strains are considered ideal for the fermentation industry (Yao et al., 2021). SBM fermented by Bacillus yields products with high nutritional value and low levels of antinutritional factors (ANFs), stimulating research in this area (Yuan et al., 2017). Kumari et al. (2023) demonstrated that fermentation with Bacillus species effectively degrades antigen proteins and increases crude protein content. The degradation of antigen proteins relies on protease hydrolysis. Low protease production is the major obstacle hindering the widespread use of microbial fermentation techniques.
Bacillus amyloliquefaciens/metabolism* ; Fermentation ; Glycine max/metabolism* ; Soybean Proteins/metabolism* ; Peptide Hydrolases/metabolism* ; Ribosomes/metabolism* ; Globulins ; Antigens, Plant ; Seed Storage Proteins

OBJECTIVE: Benzylisoquinoline alkaloids (BIAs) have pharmacological functions and clinical use. BIAs are mainly distributed in plant species across the order Ranunculales and the genus Phellodendron from Sapindales. The BIA biosynthesis has been intensively investigated in Ranunculales species. However, the accumulation mechanism of BIAs in Phellodendron is largely unknown. The aim of this study is to unravel the biosynthetic pathways of BIAs in Phellodendron amurens. METHODS: The transcriptome and metabolome data from 18 different tissues of P. amurense were meticulously sequenced and subsequently subjected to a thorough analysis. Weighted gene co-expression network analysis (WGCNA), a powerful systems biology approach that facilitates the construction and subsequent analysis of co-expression networks, was utilized to identify candidate genes involved in BIAs biosynthesis. Following this, recombinant plasmids containing candidate genes were expressed in Escherichia coli, a widely used prokaryotic expression system. The purpose of this genetic engineering endeavor was to express the candidate genes within the bacteria, thereby enabling the assessment of the resultant enzyme activity. RESULTS: The synonymous substitutions per synonymous site for paralogs indicated that at least one whole genome duplication event has occurred. The potential BIA biosynthetic pathway of P. amurense was proposed, and two PR10/Bet v1 members, 14 CYP450s, and 33 methyltransferases were selected as related to BIA biosynthesis. One PR10/Bet v1 was identified as norcoclaurine synthase, which could catalyze dopamine and 4-hydroxyphenylacetaldehyde into (S)-norcoclaurine. CONCLUSION Our studies provide important insights into the biosynthesis and evolution of BIAs in non-Ranunculales species.

OBJECTIVES: This study aims to analyze the clinical symptoms and imaging manifestations in patients with temporomandibular disorder syndrome (TMD), who are sensitive to sudden temperature drop. METHODS: One hundred and nineteen patients with TMD who attended the Department of Stomatology of the First Medical Center of Chinese People's Liberation Army General Hospital from December 2022 to December 2023 were included, including 44 males and 75 females, with a mean age of 32.4±13.7 years.The questionnaire was used to determine whether they were sensitive to temperature drop, and the TMD patients were divided into a temperature plunge-sensitive group and a temperature drop insensitive group. The clinical symptoms and imaging manifestations of patients in the two groups were observed. SPSS 25.0 was used for statistical analysis. RESULTS: There was no statistically significant difference between the gender and age of patients in the temperature plunge-sensitive group (50 patients) and the insensitivity group (69 patients) (P>0.05). The percentage of patients with pain was slightly higher in the temperature plunge-sensitive group [86.0% (43/50)] than in the insensitive group [68.1% (47/69)], and the difference was statistically significant (χ²=5.031, P=0.025), while the differences in joint murmur and mouth opening limitation between the two groups were not statistically significant. A total of 238 lateral joints were detected in both groups, the percentage of osteoarthropathic imaging changes was significantly higher in the temperature plunge-sensitive group [82.0% (82/100)] than in the insensitive group [53.6% (74/138)] (χ²=20.675, P<0.001). Magnetic imaging showed that the percentage of joint effusion was higher in patients in the temperature plunge-sensitive group [66.0% (33/50)] than in the insensitive group [42.0% (29/69)], and the difference was statistically significant (χ²=5.602, P=0.018). CONCLUSIONS TMD patients with maxillofacial pain symptoms, joint effusions, and abnormal imaging of osteoarticular structures are more likely to be sensitive to sudden temperature drops.
Humans ; Male ; Female ; Adult ; Temporomandibular Joint Disorders/diagnosis* ; Surveys and Questionnaires ; Middle Aged ; Young Adult ; Temperature ; Adolescent

OBJECTIVES: This study aimed to investigate the correlation between clinical symptoms and unilateral chewing habits in patients with temporomandibular disorder (TMD) accompanied by tinnitus. METHODS: A total of 285 patients diagnosed with TMD at the Department of Stomatology of the First Medical Center of Chinese People's Liberation Army General Hospital between December 2020 and May 2024 were included and divided into two groups: tinnitus group and non-tinnitus group. Analysis was conducted on the proportion of patients with unilateral chewing habits in both groups, the correlation between the side of tinnitus and the side of unilateral chewing, and the correlation of tinnitus with TMD clinical symptoms (joint clicking, joint pain, and limited mouth opening) and unilateral chewing habits. The correlation of the type of disc displacement with unilateral chewing and tinnitus was also examined. RESULTS: In the tinnitus group, the proportions of patients with and without unilateral chewing habits were 90.70% (39/43) and 9.30% (4/43), respectively. In the non-tinnitus group, the proportions of patients with and without unilateral chewing habits were 76.03% (184/242) and 23.97% (58/242), respectively. The proportion of patients with unilateral chewing habits in the tinnitus group was significantly higher than in the non-tinnitus group (χ²=4.613, P<0.05). Correlation analysis showed a positive correlation between tinnitus and unilateral chewing habits (P<0.05). In the left-sided tinnitus group, the proportion of left-sided unilateral chewers [54.55% (12/22)] was higher than that of right-sided unilateral chewers [45.45% (10/22)]. In the right-sided tinnitus group, the proportion of right-sided unilateral chewers [81.82% (9/11)] was higher than that of left-sided unilateral chewers [18.18% (2/11)]. The difference was statistically significant (χ²=7.282, P<0.05). A positive correlation was also found between the side of tinnitus and the side of unilateral chewing habits (P<0.05). The proportion of patients with pain was significantly higher in the tinnitus group than in the non-tinnitus group (P<0.05). No significant difference in the proportion of joint clicking or limited mouth opening and disc displacement (no disc displacement, unilateral disc displacement, bilateral disc displacement, reducible disc displacement, or irreducible disc displacement) was found between the tinnitus and non-tinnitus groups (P>0.05). CONCLUSIONS TMD with unilateral chewing habits may be a contributing factor to unexplained tinnitus. Unexplained tinnitus is correlated with joint pain in patients with TMD.
Humans ; Tinnitus/physiopathology* ; Temporomandibular Joint Disorders/physiopathology* ; Mastication ; Male ; Adult ; Female ; Middle Aged ; Habits

Foot-and-mouth disease (FMD) is one of the major animal infectious diseases in the world. All cloven-hoofed animals are susceptible to FMD. Vaccination is still the first choice for the prevention and control of FMD. mRNA vaccines can be rapidly designed, synthesized, and produced on a large scale in vitro, and they can induce effective protective immune responses, demonstrating the advantages of rapid development, easy preparation, and low biosafety risks. The design of untranslated regions is a key to enhancing the expression and efficacy of mRNA vaccines. In order to generate an efficient FMD mRNA vaccine, we designed three FMD P12A3C expression vectors with different untranslated regions and synthesized corresponding mRNAs. By comparing expression efficiency of these vectors and their mRNAs at different time points and in different cell lines, we found that the mRNA P12A3C-UTR3 had the best expression and universality. This study laid a foundation for the development of mRNA vaccines against FMD and provided a theoretical basis for the optimal sequence design of efficient mRNA.
Foot-and-Mouth Disease Virus/genetics* ; Animals ; RNA, Messenger/biosynthesis* ; Foot-and-Mouth Disease/immunology* ; Antigens, Viral/biosynthesis* ; Viral Vaccines/biosynthesis* ; Genetic Vectors/genetics* ; Cell Line ; Vaccines, DNA/immunology*