AIM To study the chemical constituents from salt-processed Citri Reticulatae Semen and their antioxidant activities.METHODS The 85% ethanol extract from salt-processed Citri Reticulatae Semen was isolated and purified by silica gel,D101 macroporous resins,MCI,ODS,Sephadex LH-20 and semi-prepative HPLC,then the structures of obtained compounds were identified by physicochemical properties and spectral data.Their antioxidant activities in vitro of the ethanol extract of Citri Reticulatae Semen,salt-processed Citri Reticulatae Semen and all the obtained compounds were evaluated by DPPH and ABTS+assay.RESULTS Sixteen compounds were isolated and identified as limonin(1),obacunone(2),nomilin(3),deacetyl nomilin(4),kaempferol(5),nobiletin(6),diosmetin(7),isosakuranetin(8),hesperetin(9),epicatechin(10),trans-p-menthane-1α,2β,8-triol(11),byakangelicin(12),vanillin(13),p-coumaric acid(14),4-[(1-ethoxy-2-hydroxy)ethyl]phenol(15),catechol(16).Compound 10 showed strong DPPH free radical scavenging activity,with an IC50 value of(0.015±0.001)μmol/mL,and strong ABTS+radical scavenging activity,with an IC50 value of(0.010±0.005)μmol/mL.CONCLUSION Compounds 8,11,15-16 are isolated from genus Citrus for the first time,5,12,14 are obtained from Citri Reticulatae Semen for the first time.Compound 10 shows obvious antioxidant activities.After salt roasting,the antioxidant activity of the ethanol extract of Citri Reticulatae Semen is enhanced.

AIM To study the chemical constituents from the leaves of Citrus reticulata Blanco and their anti-inflammatory activities.METHODS The 85%ethanol extract from the leaves of C.reticulata was isolated and purified by silica gel,D101 macroporous resin,MCI,ODS and Sephadex LH-20,then the structures of obtained compounds were identified by physicochemical properties and spectral data.The Griess method was used to determine their inhibitory activities on lipopolysaccharide-induced NO production in macrophages RAW 264.7 cells.The mice foot swelling inflammation model induced by carrageenan was established,and the levels of IL-1β,TNF-α were detected.RESULTS Twelve compounds were isolated and identified as nobiletin(1),tangeretin(2),5-demethylinoblitin(3),5,4'-dihydroxy-6,7,8,3'-tetramethoxy flavone(4),5-hydroxy-7,8,3',4'-trimethoxyflavanone(5),3,5,6,7,8,3',4'-heptamethoxyflavanone(6),hesperetin(7),5-hydroxy-6,7,3',4'-tetramethoxyphenone(8),β-balsam alcohol(9),stigmaster-5-en-3β-alcohol(10),p-hydroxybenzaldehyde(11),vanillin(12).Compounds 1,4,6,7,10 and 12 had strong inhibitory activites on NO release in LPS-induced RAW 264.7 cells,and the IC50 values were(25.21±2.10),(37.77±0.50),(38.19±1.58),(21.89±1.73),(43.81±1.18),(47.98±2.55),(41.23±1.11),(43.80±1.43)μmol/mL,respectively.Compounds 2-3 reduced IL-1β and TNF-α levels(P＜0.05,P＜0.01).CONCLUSION Compounds 6-7,9 are isolated from this plant for the first time.Compounds 1-4,8 exhibit strong in vitro anti-inflammatory activities,and compounds 2-3 exhibit significant in vivo anti-inflammatory activities.

Objective To investigate the incidence rate,clinical characteristics,and prognosis of neonatal stroke in Shenzhen,China.Methods Led by Shenzhen Children's Hospital,the Shenzhen Neonatal Data Collaboration Network organized 21 institutions to collect 36 cases of neonatal stroke from January 2020 to December 2022.The incidence,clinical characteristics,treatment,and prognosis of neonatal stroke in Shenzhen were analyzed.Results The incidence rate of neonatal stroke in 21 hospitals from 2020 to 2022 was 1/15 137,1/6 060,and 1/7 704,respectively.Ischemic stroke accounted for 75％(27/36);boys accounted for 64％(23/36).Among the 36 neonates,31(86％)had disease onset within 3 days after birth,and 19(53％)had convulsion as the initial presentation.Cerebral MRI showed that 22 neonates(61％)had left cerebral infarction and 13(36％)had basal ganglia infarction.Magnetic resonance angiography was performed for 12 neonates,among whom 9(75％)had involvement of the middle cerebral artery.Electroencephalography was performed for 29 neonates,with sharp waves in 21 neonates(72％)and seizures in 10 neonates(34％).Symptomatic/supportive treatment varied across different hospitals.Neonatal Behavioral Neurological Assessment was performed for 12 neonates(33％,12/36),with a mean score of(32±4)points.The prognosis of 27 neonates was followed up to around 12 months of age,with 44％(12/27)of the neonates having a good prognosis.Conclusions Ischemic stroke is the main type of neonatal stroke,often with convulsions as the initial presentation,involvement of the middle cerebral artery,sharp waves on electroencephalography,and a relatively low neurodevelopment score.Symptomatic/supportive treatment is the main treatment method,and some neonates tend to have a poor prognosis.

Purpose::Vibrio vulnificus ( V. Vulnificus) infection is characterized by rapid onset, aggressive progression, and challenging treatment. Bacterial resistance poses a significant challenge for clinical anti-infection treatment and is thus the subject of research. Enhancing host infection tolerance represents a novel infection prevention strategy to improve patient survival. Our team initially identified cytochrome P4501A1 (CYP1A1) as an important target owing to its negative modulation of the body's infection tolerance. This study explored the superior effects of the CYP1A1 inhibitor bergamottin compared to antibiotic combination therapy on the survival of mice infected with multidrug-resistant V. Vulnificus and the protection of their vital organs. Methods::An increasing concentration gradient method was used to induce multidrug-resistant V. Vulnificus development. We established a lethal infection model in C57BL/6J male mice and evaluated the effect of bergamottin on mouse survival. A mild infection model was established in C57BL/6J male mice, and the serum levels of creatinine, urea nitrogen, aspartate aminotransferase, and alanine aminotransferase were determined using enzyme-linked immunosorbent assay to evaluate the effect of bergamottin on liver and kidney function. The morphological changes induced in the presence of bergamottin in mouse organs were evaluated by hematoxylin and eosin staining of liver and kidney tissues. The bacterial growth curve and organ load determination were used to evaluate whether bergamottin has a direct antibacterial effect on multidrug-resistant V. Vulnificus. Quantification of inflammatory factors in serum by enzyme-linked immunosorbent assay and the expression levels of inflammatory factors in liver and kidney tissues by real-time quantitative polymerase chain reaction were performed to evaluate the effect of bergamottin on inflammatory factor levels. Western blot analysis of IκBα, phosphorylated IκBα, p65, and phosphorylated p65 protein expression in liver and kidney tissues and in human hepatocellular carcinomas-2 and human kidney-2 cell lines was used to evaluate the effect of bergamottin on the nuclear factor kappa-B signaling pathway. One-way ANOVA and Kaplan-Meier analysis were used for statistical analysis. Results::In mice infected with multidrug-resistant V. Vulnificus, bergamottin prolonged survival ( p = 0.014), reduced the serum creatinine ( p = 0.002), urea nitrogen ( p = 0.030), aspartate aminotransferase ( p = 0.029), and alanine aminotransferase ( p = 0.003) levels, and protected the cellular morphology of liver and kidney tissues. Bergamottin inhibited interleukin (IL)-1β, IL-6, and tumor necrosis factor (TNF)-α expression in serum (IL-1β: p = 0.010, IL-6: p = 0.029, TNF-α: p = 0.025) and inhibited the protein expression of the inflammatory factors IL-1β, IL-6, TNF-α in liver (IL-1β: p = 0.010, IL-6: p = 0.011, TNF-α: p = 0.037) and kidney (IL-1β: p = 0.016, IL-6: p = 0.011, TNF-α: p = 0.008) tissues. Bergamottin did not affect the proliferation of multidrug-resistant V. Vulnificus or the bacterial load in the mouse peritoneal lavage fluid ( p = 0.225), liver ( p = 0.186), or kidney ( p = 0.637). Conclusion::Bergamottin enhances the tolerance of mice to multidrug-resistant V. Vulnificus infection. This study can serve as a reference and guide the development of novel clinical treatment strategies for V. Vulnificus.

ObjectiveTo investigate the expression of RNA binding motif single stranded interacting protein 3 （RBMS3） in epithelial ovarian cancer （EOC） tissues and its relationship with the clinicopathological features and prognosis of EOC. MethodsThe study enrolled the paraffin-embedded tissues from 110 EOC cases and 73 benign epithelial ovarian tumor cases pathologically diagnosed in the first affiliated Hospital of Bengbu Medical College from January 2015 to December 2019. By using anti-RBMS3 polyclonal antibody， the immunohistochemical staining was employed to detect RBMS3 expression in the tissues and then its correlation with the clinicopathological parameters and prognosis of EOC was analyzed. ResultsRBMS3 was expressed in both EOC and benign epithelial ovarian tumor tissues. RBMS3 expression in EOC tissues， significantly related with International Federation of Gynecology and Obstetrics （FIGO） stage， histological grade， CEA levels and survival status， was significantly lower than that in benign epithelial ovarian tumor tissues （P<0.05）. Kaplan–Meier survival curve showed that low RBMS3 expression in EOC patients was correlated with decreased progression-free survival （PFS） and overall survival （OS） （P<0.05）. Univariate analysis showed that RBMS3 expression， FIGO stage， residual lesion size， intestinal metastasis and intraperitoneal implantation were associated with OS of EOC patients （P<0.05）； multivariate analysis showed that low RBMS3 expression and intestinal metastasis were independent risk factors for poor prognosis in EOC patients （P<0.05）. ConclusionsRBMS3 is expressed at low levels in EOC tissues， which is closely related to poor prognosis of EOC patients. RBMS3 may function as a tumor suppressor gene in EOC tissues and can be used as an EOC-independent prognostic marker for targeted therapy against EOC.

OBJECTIVE: To delineate the onset and recurrence characteristics of noncardiogenic ischemic stroke patients in China. METHODS: A prospective, multicenter and registry study was carried out in 2,558 patients at 7 representative clinical sub-centers during November 3, 2016 to February 17, 2019. A questionnaire was used to collect information of patients regarding CM syndromes and constitutions and associated risk factors. Additionally, stroke recurrence was defined as a primary outcome indicator. RESULTS: A total of 327 (12.78 %) patients endured recurrence events, 1,681 (65.72%) were men, and the average age was 63.33 ± 9.45 years. Totally 1,741 (68.06%) patients suffered first-ever ischemic stroke, 1,772 (69.27%) patients reported to have hypertension, and 1,640 (64.11%) of them reported dyslipidemia, 1,595 (62.35%) patients exhibited small-artery occlusion by The Trial of Org 10172 in Acute Stroke Treatment (TOAST) classification. Specifically, 1,271 (49.69%) patients were considered as qi-deficient constitution, and 1,227 (47.97%) patients were determined as stagnant blood constitution. There were 1,303 (50.94%) patients diagnosed as blood stasis syndrome, 1,280 (50.04%) patients exhibited phlegm and dampness syndrome and 1,012 (39.56%) patients demonstrated qi deficiency syndrome. And 1,033 (40.38%) patients declared intracranial artery stenosis, and 478 (18.69%) patients reported carotid artery stenosis. The plaque in 1,508 (41.36%) patients were of mixed. Particularly, 41.09% of them demonstrated abnormal levels of glycated hemoglobin levels. CONCLUSIONS Recurrence in minor and small-artery stroke cannot be ignored. Hypertension, dyslipidemia, abnormal HbA1c, intracranial artery stenosis and carotid plaque were more common in stroke patients. Particularly, phlegm-dampness and blood stasis syndromes, as well as qi deficiency and blood stasis constitutions, were still the main manifestations of stroke. (Trial registration at ClinicalTrials.gov No. NCT03174535).
Aged ; Constriction, Pathologic ; Female ; Hospitals ; Humans ; Hypertension ; Ischemic Stroke ; Male ; Medicine, Chinese Traditional ; Middle Aged ; Prospective Studies ; Stroke/epidemiology* ; Syndrome

Chinese Pharmacopoeia is an important part of drug standards in China, and it is also a legal basis that must be strictly followed in drug development, production, operation, application, and management. The information on prescriptions, preparation methods, properties, identification, inspection, content determination, functions and indications, usage and dosage, precautions, specifications, and storage of Chinese patent medicine preparations included in the Chinese Pharmacopoeia(Vol.Ⅰ) was clarified. The "Preparation Method" section describes the preparation process of Chinese patent medicine from decoction pieces to finished preparations in detail and specifies the preparation production methods and parameters, which has a good guiding and standardizing effect on the production of Chinese patent medicine in China. The present study summarized the preparation methods of Chinese patent medicine preparations and single drug preparations contained in the Chinese Pharmacopoeia(2020 edition, Vol.Ⅰ) in stages and analyzed the common preparation methods and technical parameters of Chinese patent medicine preparations, which is helpful to understand the current situation of Chinese patent medicine production technology in China and can provide references for the development of new Chinese medicine, the transformation of large varieties of Chinese patent medicine, and the optimization of preparation process of Chinese patent medicine in the market.
China ; Drugs, Chinese Herbal ; Medicine, Chinese Traditional ; Nonprescription Drugs ; Prescriptions

Objective: To explore the effects of porcine acellular dermal matrix (ADM) combined with human epidermal stem cells (ESCs) on wound healing of full-thickness skin defect in nude mice. Methods: The morphology of porcine ADM was analyzed by photograph of digital camera, the cell residues in porcine ADM were observed by hematoxylin-eosin (HE) staining, the surface structure of porcine ADM was observed by scanning electron microscope, the secondary structure of porcine ADM was analyzed by infrared spectrometer, the porcine ADM particle size was analyzed by dynamic light scattering particle size analyzer, and the porcine ADM potential was analyzed by nano-particle size potentiometer. The morphology of porcine ADM was observed by inverted fluorescence microscope when it was placed in culture medium for 30 min, 1 d, and 5 d (n=2). The porcine ADM was divided into 5 min group, 10 min group, 20 min group, 30 min group, 60 min group, and 120 min group according to the random number table (the same grouping method below) in static state at normal temperature for the corresponding time to calculate the water absorption by weighing method (n=3). Swiss white mouse embryonic fibroblasts (Fbs) were divided into blank control group (culture medium only), and 50.0 g/L ADM extract group, 37.5 g/L ADM extract group, 25.0 g/L ADM extract group, 12.5 g/L ADM extract group, and 6.5 g/L ADM extract group which were added with the corresponding final concentrations of ADM extract respectively. At post culture hour (PCH) 24, 48, and 72, the cell survival rate was detected by cell counting kit 8 and the cytotoxicity was graded (n=5). The erythrocytes of a 6-week-old male Sprague-Dawley male rat were divided into normal saline group, ultra-pure water group, and 5 mg/mL ADM extract group, 10 mg/mL ADM extract group, and 15 mg/mL ADM extract group which were treated with the corresponding final concentrations of porcine ADM extract respectively. After reaction for 3 h, the absorbance value of hemoglobin was detected by microplate reader to represent the blood compatibility of porcine ADM (n=3). ESCs were isolated and cultured from the discarded prepuce of a 6-year-old healthy boy who was treated in the Department of Urology of the First Affiliated Hospital of Army Medical University (the Third Military Medical University) in July 2020, and then identified by flow cytometry. The porcine ADM particles of composite ESC (hereinafter referred to as ESC/ADM) were constructed by mixed culture. After 3 days of culture, the composite effect of ESC/ADM was observed by HE staining and laser scanning confocal microscope. Thirty-six 7-8-week-old male non-thymic nude mice were divided into phosphate buffer solution (PBS) alone group, ADM alone group, ESC alone group, and ESC/ADM group, with 9 mice in each group, and the wound model of full-thickness skin defect was established. Immediately after injury, the wounds were treated with the corresponding reagents at one time. On post injury day (PID) 1, 7, 11, and 15, the wound healing was observed and the wound healing rate was counted (n=3). On PID 7, the epithelialization of wounds was observed by HE staining and the length of un-epithelialized wound was measured (with this and the following sample numbers of 4). On PID 11, the dermal area and collagen deposition of wounds were observed by Masson staining and the dermal area of wound section was calculated, the number of cells expressing CD49f, a specific marker of ESC, was calculated with immunofluorescence staining, the mRNA expression of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) in ESC after wound transplantation was detected by real-time fluorescence quantitative reverse transcription polymerase chain reaction. Data were statistically analyzed with independent sample t test, one-way analysis of variance, analysis of variance for repeated measurement, and least significant difference t test. Results: The porcine ADM was white particles and composed of reticular structure, with no cells inside, disordered structure, and rough surface. The absorption peak of porcine ADM appeared at the wave numbers of 1 659, 1 549, and 1 239 cm^-1, respectively. The main particle size distribution of porcine ADM in solution was 500 to 700 nm, with negative charge on the surface. The morphology of porcine ADM in static state at 30 min and on 1 and 5 d was relatively stable. The water absorption of porcine ADM remained relatively high level in static state from 30 min to 120 min. The cytotoxicity of mouse embryonic Fbs in 6.5 g/L ADM extract group, 12.5 g/L ADM extract group, and 25.0 g/L ADM extract group was grade 1 at PCH 24, and the cytotoxicity of the other groups was 0 grade at each time point. After reaction for 3 h, the absorbance value of hemoglobin of erythrocytes in ultra-pure water group was significantly higher than the values in normal saline group and 15 mg/mL ADM extract group (with t values of 8.14 and 7.96, respectively, P<0.01). After 3 days of culture, the cells of the fourth passage showed pebble-like morphology, with low expression of CD71 and high expression of CD49f, which were identified as ESCs. There was ESC attachment and growth on porcine ADM particles. On PID 1, the wound sizes of nude mice were almost the same in PBS alone group, ADM alone group, ESC alone group, and ESC/ADM group. On PID 7, 11, and 15, the wound contraction of nude mice in each group was observed, especially in ADM alone group, ESC alone group, and ESC/ADM group. On PID 7, the wound healing rates of nude mice in ESC alone group and ESC/ADM group were significantly higher than the rate in PBS alone group (with t values of 2.83 and 4.72 respectively, P<0.05 or P<0.01). On PID 11, the wound healing rate of nude mice in ESC/ADM group was significantly higher than that in PBS alone group (t=4.86, P<0.01). On PID 15, the wound healing rates of nude mice in ADM alone group, ESC alone group, and ESC/ADM group were significantly higher than the rate in PBS alone group (with t values of 2.71, 2.90, and 3.23 respectively, P<0.05). On PID 7, the length of un-epithelialized wound of nude mice in ADM alone group, ESC alone group, and ESC/ADM group was (816±85), (635±66), and (163±32) μm, respectively, which were significantly shorter than (1 199±43) μm in PBS alone group (with t values of 5.69, 10.19, and 27.54 respectively, P<0.01). On PID 11, the dermal areas of wound section of nude mice in ADM alone group, ESC alone group, and ESC/ADM group were significantly larger than the area in PBS alone group (with t values of 27.14, 5.29, and 15.90 respectively, P<0.01); the collagen production of nude mice in ADM alone group and ESC/ADM group was more obvious than that in PBS alone group, and the collagen production of nude mice in ESC alone group and PBS alone group was similar. On PID 11, in the wounds of nude mice in ESC alone group and ESC/ADM group, the cells with positive expression of CD49f were respectively 135±7 and 185±15, and the mRNA expressions of GAPDH were positive; while there were no expressions of CD49f nor mRNA of GAPDH in the wounds of nude mice in PBS alone group and ADM alone group. Conclusions: ESC/ADM particles can promote the wound healing of full-thickness skin defects in nude mice, which may be related to the improved survival rate of ESCs after transplantation and the promotion of dermal structure rearrangement and angiogenesis by ADM.
Acellular Dermis ; Animals ; Fibroblasts ; Humans ; Male ; Mice ; Mice, Nude ; Rats ; Rats, Sprague-Dawley ; Stem Cells ; Swine ; Wound Healing

BACKGROUND: The prevalence of skin diseases and diabetes mellitus (DM) are prominent around the world. The current scope of knowledge regarding the prevalence of skin diseases and comorbidities with type 2 DM (T2DM) is limited, leading to limited recognition of the correlations between skin diseases and T2DM. METHODS: We collected 383 subjects from the Da Qing Diabetes Study during the period from July 9th to September 1st, 2016. The subjects were categorized into three groups: Normal glucose tolerance (NGT), impaired glucose tolerance (IGT), and T2DM. The prevalence and clinical characteristics of skin diseases were recorded and investigated. RESULTS: In this cross-sectional study, 383 individuals with ages ranging from 53 to 89-year-old were recruited. The overall prevalence of skin diseases was 93.5%, and 75.7% of individuals had two or more kinds of skin diseases. Additionally, there were 47 kinds of comorbid skin diseases in patients with T2DM, of which eight kinds of skin diseases had a prevalence >10%. The prevalence of skin diseases in NGT, IGT, and T2DM groups were 93.3%, 91.5%, and 96.6%, respectively; stratified analysis by categories showed a statistically significant difference in "disturbances of pigmentation" and "neurological and psychogenic dermatoses". The duration of T2DM also significantly associated with the prevalence of "disturbances of pigmentation" and "neurological and psychogenic dermatoses". Subsequently, the prevalence of "disturbances of pigmentation" was higher in males than females in NGT (P < 0.01) and T2DM (P < 0.01) groups. In addition, the difference in the prevalence of "disturbances of pigmentation" was also significant in NGT and T2DM groups (P < 0.01). CONCLUSIONS There was a high prevalence of skin diseases in the Da Qing Diabetes Study. To address the skin diseases in the Da Qing Diabetes Study, increased awareness and intervention measures should be implemented.
Aged ; Aged, 80 and over ; Blood Glucose ; Cross-Sectional Studies ; Diabetes Mellitus, Type 2/epidemiology* ; Female ; Glucose Intolerance/epidemiology* ; Glucose Tolerance Test ; Humans ; Male ; Middle Aged ; Skin Diseases/epidemiology*

Aim To explore the effect of THPA1 in the metasta- sis of gastric cancer and the underlying mechanism. Methods The correlation between TRPA1 and the survival time of gastric cancer patients was analyzed using Kaplan-Meier plotter data base. The expressions of TRPA1 in different cells were detected by Western blot. Docking was used to explore the binding poten tial between cardamonin and TRPA1. Long-term dynamic cell imaging, CCK-8 and Transwell were used to evaluate the effects of HC-030031 and cardamonin on the proliferation and migration of MKN-45 cells. The differential metabolites between normal gastric epithelial cells and gastric cancer cells were studied by GC-MS. Results The expression of TRPA1 in gastric cancer patients was significantly negatively correlated with their surviv al. TRPA1 was overexpressed in gastric cancer cells. And the migration of gastric cancer cells was positively correlated with the expression and activation of TRPA1. Cardamonin had similar pharmacological effects with HC-030031, both of which could reduce the migration of gastric cancer cells. The metabolic path ways of asparagine and myo-inositol were found to be different between gastric cancer cells and normal gastric epithelial cells by cell metabolomics analysis. Conclusions TRPAI may be an indicator for detecting gastric cancer metastasis. Cardamonin in hibits metastasis by binding to TRPAI, meanwhile restrains the activation of TRPAI. Cardamonin may inhibit the function of TRPAI by binding to TRPAI, playing a role in inhibiting gastric cancer metastasis.