OBJECTIVE: Gastric cancer (GC) is one of the most common malignancies seen in clinic and requires novel treatment options. Morin is a natural flavonoid extracted from the flower stalk of a highly valuable medicinal plant Prunella vulgaris L., which exhibits an anti-cancer effect in multiple types of tumors. However, the therapeutic effect and underlying mechanism of morin in treating GC remains elusive. The study aims to explore the therapeutic effect and underlying molecular mechanisms of morin in GC. METHODS: For in vitro experiments, the proliferation inhibition of morin was measured by cell counting kit-8 assay and colony formation assay in human GC cell line MKN45, human gastric adenocarcinoma cell line AGS, and human gastric epithelial cell line GES-1; for apoptosis analysis, microscopic photography, Western blotting, ubiquitination analysis, quantitative polymerase chain reaction analysis, flow cytometry, and RNA interference technology were employed. For in vivo studies, immunohistochemistry, biomedical analysis, and Western blotting were used to assess the efficacy and safety of morin in a xenograft mouse model of GC. RESULTS: Morin significantly inhibited the proliferation of GC cells MKN45 and AGS in a dose- and time-dependent manner, but did not inhibit human gastric epithelial cells GES-1. Only the caspase inhibitor Z-VAD-FMK was able to significantly reverse the inhibition of proliferation by morin in both GC cells, suggesting that apoptosis was the main type of cell death during the treatment. Morin induced intrinsic apoptosis in a dose-dependent manner in GC cells, which mainly relied on B cell leukemia/lymphoma 2 (BCL-2) associated agonist of cell death (BAD) but not phorbol-12-myristate-13-acetate-induced protein 1. The upregulation of BAD by morin was due to blocking the ubiquitination degradation of BAD, rather than the transcription regulation and the phosphorylation of BAD. Furthermore, the combination of morin and BCL-2 inhibitor navitoclax (also known as ABT-737) produced a synergistic inhibitory effect in GC cells through amplifying apoptotic signals. In addition, morin treatment significantly suppressed the growth of GC in vivo by upregulating BAD and the subsequent activation of its downstream apoptosis pathway. CONCLUSION Morin suppressed GC by inducing apoptosis, which was mainly due to blocking the ubiquitination-based degradation of the pro-apoptotic protein BAD. The combination of morin and the BCL-2 inhibitor ABT-737 synergistically amplified apoptotic signals in GC cells, which may overcome the drug resistance of the BCL-2 inhibitor. These findings indicated that morin was a potent and promising agent for GC treatment. Please cite this article as: Wang Y, Sun XY, Ma FQ, Ren MM, Zhao RH, Qin MM, Zhu XH, Xu Y, Cao ND, Chen YY, Dong TG, Pan YF, Zhao AG. Morin inhibits ubiquitination degradation of BCL-2 associated agonist of cell death and synergizes with BCL-2 inhibitor in gastric cancer cells. J Integr Med. 2025; 23(3): 320-332.
Humans ; Flavonoids/therapeutic use* ; Stomach Neoplasms/pathology* ; Animals ; Proto-Oncogene Proteins c-bcl-2/metabolism* ; Cell Line, Tumor ; Apoptosis/drug effects* ; Cell Proliferation/drug effects* ; Ubiquitination/drug effects* ; Mice ; Drug Synergism ; Mice, Inbred BALB C ; Mice, Nude ; Xenograft Model Antitumor Assays ; Flavones

Objective: To investigate the effects of exosomes from human adipose-derived mesenchymal stem cells (ADSCs) on pulmonary vascular endothelial cells (PMVECs) injury in septic mice and its mechanism. Methods: The experimental research method was adopted. The primary ADSCs were isolated and cultured from the discarded fresh adipose tissue of 3 patients (female, 10-25 years old), who were admitted to the First Affiliated Hospital of Air Force Medical University undergoing abdominal surgery, and the cell morphology was observed by inverted phase contrast microscope on the 5^th day. The expressions of CD29, CD34, CD44, CD45, CD73, and CD90 of ADSCs in the third passage were detected by flow cytometry. The third to the fifth passage of ADSCs were collected, and their exosomes from the cell supernatant were obtained by differential ultracentrifugation, and the shape, particle size, and the protein expressions of CD9, CD63, tumor susceptibility gene 101 (TSG101), and β-actin of exosomes were detected, respectively, by transmission electron microscopy, nano-particle tracking analysis and Western blotting. Twenty-four adult male BALB/c mice were adopted and were divided into normal control group, caecal ligation perforation (CLP) alone group, and CLP+ADSC-exosome group with each group of 8 according to random number table (the same grouping method below) and were treated accordingly. At 24 h after operation, tumor necrosis factor (TNF-α) and interleukin 1β (IL-1β) levels of mice serum were detected by enzyme-linked immunosorbent assay, and lung tissue morphology of mice was detected by hematoxylin-eosin and myeloperoxidase staining, and the expression of 8-hydroxy-deoxyguanosine (8-OHdG) of mouse lung cells was detected by immunofluorescence method. Primary PMVECs were obtained from 1-month-old C57 mice regardless gender by tissue block method. The expression of CD31 of PMVECs was detected by immunofluorescence and flow cytometry. The third passage of PMVECs was co-cultured with ADSCs derived exosomes for 12 h, and the phagocytosis of exosomes by PMVECs was detected by PKH26 kit. The third passage of PMVECs were adopted and were divided into blank control group, macrophage supernatant alone group, and macrophage supernatant+ADSC-exosome group, with 3 wells in each group, which were treated accordingly. After 24 h, the content of reactive oxygen species in cells was detected by flow cytometry, the expression of 8-OHdG in cells was detected by immunofluorescence, and Transwell assay was used to determine the permeability of cell monolayer. The number of samples in above were all 3. Data were statistically analyzed with one-way analysis of variance and least significant difference t test. Results: The primary ADSCs were isolated and cultured to day 5, growing densely in a spindle shape with a typical swirl-like. The percentages of CD29, CD44, CD73 and CD90 positive cells of ADSCs in the third passage were all >90%, and the percentages of CD34 and CD45 positive cells were <5%. Exosomes derived from ADSCs of the third to fifth passages showed a typical double-cavity disc-like structure with an average particle size of 103 nm, and the protein expressions of CD9, CD63 and TSG101 of exosomes were positive, while the protein expression of β-actin of exosomes was negative. At 24 h after operation, compared with those in normal control group, both the levels of TNF-α and IL-1β of mice serum in CLP alone group were significantly increased (with t values of 28.76 and 29.69, respectively, P<0.01); compared with those in CLP alone group, both the content of TNF-α and IL-1β of mice serum in CLP+ADSC-exosome group was significantly decreased (with t values of 9.90 and 4.76, respectively, P<0.05 or P<0.01). At 24 h after surgery, the pulmonary tissue structure of mice in normal control group was clear and complete without inflammatory cell infiltration; compared with those in normal control group, the pulmonary tissue edema and inflammatory cell infiltration of mice in CLP alone group were more obvious; compared with those in CLP alone group, the pulmonary tissue edema and inflammatory cell infiltration of mice in CLP+ADSC-exosome group were significantly reduced. At 24 h after operation, endothelial cells in lung tissues of mice in 3 groups showed positive expression of CD31; compared with that in normal control group, the fluorescence intensity of 8-OHdG positive cells of the lung tissues of mice in CLP alone group was significantly increased, and compared with that in CLP alone group, the fluorescence intensity of 8-OHdG positive cells in the lung tissues of mice in CLP+ADSC-exosome group was significantly decreased. The PMVECs in the 3^rd passage showed CD31 positive expression by immunofluorescence, and the result of flow cytometry showed that CD31 positive cells accounted for 99.5%. At 12 h after co-culture, ADSC-derived exosomes were successfully phagocytose by PMVECs and entered its cytoplasm. At 12 h after culture of the third passage of PMVECs, compared with that in blank control group, the fluorescence intensity of reactive oxygen species of PMVECs in macrophage supernatant alone group was significantly increased (t=15.73, P<0.01); compared with that in macrophage supernatant alone group, the fluorescence intensity of reactive oxygen species of PMVECs in macrophage supernatant+ADSC-exosome group was significantly decreased (t=4.72, P<0.01). At 12 h after culture of the third passage of PMVECs, and the 8-OHdG positive fluorescence intensity of PMVECs in macrophage supernatant alone group was significantly increased; and compared with that in blank control group, the 8-OHdG positive fluorescence intensity of PMVECs in macrophage+ADSC-exosome supernatant group was between blank control group and macrophage supernatant alone group. At 12 h after culture of the third passage PMVECs, compared with that in blank control group, the permeability of PMVECs monolayer in macrophage supernatant alone group was significantly increased (t=6.34, P<0.01); compared with that in macrophage supernatant alone group, the permeability of PMVECs monolayer cells in macrophage supernatant+ADSC-exosome group was significantly decreased (t=2.93, P<0.05). Conclusions: Exosomes derived from ADSCs can ameliorate oxidative damage in mouse lung tissue, decrease the level of reactive oxygen species, 8-OHdG expression, and permeability of PMVECs induced by macrophage supernatant.
Animals ; Endothelial Cells/metabolism* ; Exosomes/metabolism* ; Female ; Humans ; Lung Injury/metabolism* ; Male ; Mesenchymal Stem Cells/metabolism* ; Mice ; Sepsis/pathology*

OBJECTIVE: To describe and assess the repair technique and perioperative management for cerebrospinal fluid (CSF) leak resulting from extensive anterior skull base fracture via extradural anterior skull base approach. METHODS: This was a retrospective review conducted at the Department of Neurosurgery of the Shanghai Tenth People's Hospital from January 2015 to April 2020. Patients with traumatic CSF rhinorrhea resulting from extensive anterior skull base fracture treated surgically via extended extradural anterior skull base approach were included in this study. The data of medical and radiological records, surgical approaches, repair techniques, peritoperative management, surgical outcome and postoperative follow-up were analyzed. Surgical repair techniques were tailored to the condition of associated injuries of the scalp, bony and dura injuries and associated intracranial lesions. Patients were followed up for the outcome of CSF leak and surgical complications. Data were presented as frequency and percent. RESULTS: Thirty-five patients were included in this series. The patients' mean age was 33 years (range 11-71 years). Eight patients were treated surgically within 2 weeks; while the other 27 patients, with prolonged or recurrent CSF rhinorrhea, received the repair surgery at 17 days to 10 years after the initial trauma. The mean overall length of follow-up was 23 months (range 3-65 months). All the patients suffered from frontobasal multiple fractures. The basic repair tenet was to achieve watertight seal of the dura. The frontal pericranial flap alone was used in 20 patients, combined with temporalis muscle and/or its facia in 10 patients. Free fascia lata graft was used instead in the rest 5 patients. No CSF leak was found in all the patients at discharge. There was no surgical mortality in this series. Bilateral anosmia was the most common complication. At follow-up, no recurrent CSF leak or meningitis occurred. No patients developed mucoceles, epidural abscess or osteomyelitis. One patient ultimately required ventriculoperitoneal shunt because of progressive hydrocephalus. CONCLUSION Traumatic CSF rhinorrhea associated with extensive anterior skull base fractures often requires aggressive treatment via extended intracranial extradural approach. Vascularized tissue flaps are ideal grafts for cranial base reconstruction, either alone or in combination with temporalis muscle and its fascia---fascia lata sometimes can be opted as free autologous graft. The approach is usually reserved for patients with traumatic CSF rhinorrhea in complex frontobasal injuries.

Objective: To investigate the safety and efficacy of enhanced recovery after surgery (ERAS) in the clinical management of hypopharyngeal squamous cell carcinoma (HSCC). Methods: In this retrospective study, a total of 168 patients with pyriform sinus carcinoma in Qilu Hospital of Shandong University from January 2015 to January 2019 were divided into two groups, based on the different perioperative interventions that patients received, i.e. the ERAS group (n=64) and the conventional group (n=104), including 164 males and 4 females, whose ages ranged from 42 to 84 years old. The difference between two groups in the operative time, postoperative nutritional status, incidences of postoperative complications and postoperative hospitalization time were compared using the student's t test, Chi-squared test or Fisher's exact test. Results: Compared with the conventional group, patients in the ERAS group had significantly shorter operative time [(166.8±58.2) min vs. (183.3±39.9) min,t=-2.72, P=0.031], higher levels of postoperative serum albumin [(38.3±4.2) μmol/L vs. (36.6±3.3) μmol/L, t=2.73, P=0.007] and more body weight [(65.4±9.4) kg vs. (62.1±9.4) kg, t=2.22, P=0.028], lower incidences of postoperative subcutaneous infection [7.8% (5/64) vs. 20.2% (21/104), χ²=4.64, P=0.03] and severe pneumonia [4.7% (3/64) vs. 15.4% (16/104), χ²=4.52, P=0.03], and shorter postoperative hospitalization time [(16.5±3.9) d vs. (18.2±4.3) d, t=-2.65, P<0.05]. Conclusion: ERAS is effective and safe in the surgical management of HSCC, with benefits in reducing the operative stress via saving operation time, shortening the hospitalization time, ameliorating nutritional status and decreasing the incidences of complications.
Adult ; Aged ; Aged, 80 and over ; Enhanced Recovery After Surgery ; Female ; Head and Neck Neoplasms ; Humans ; Length of Stay ; Male ; Middle Aged ; Postoperative Complications/epidemiology* ; Retrospective Studies ; Squamous Cell Carcinoma of Head and Neck ; Treatment Outcome

Objective: To compare the chemical components in raw products and characteristic processed products with porcine cardiac blood of Salviae Miltiorrhizae Radix et Rhizoma from Menghe medical school.Method: The ultra performance liquid chromatography-quadrupole-time of flight mass spectrometry (UPLC-Q-TOF/MS) was performed on an ACQUITY UPLC^® C₁₈ column (2.1 mm×100 mm,1.7 μm),mobile phase was 0.1% formic acid aqueous solution (A)-0.1% formic acid acetonitrile solution (B) for gradient elution (0-1 min,8% B,1-1.5 min,8%-20% B,1.5-4 min,20% B,4-5 min,20%-60% B,5-9 min,60%-70% B,9-10 min,70%-95% B,10-13 min,95% B).The column temperature was 40℃ and the flow rate was 0.3 mL·min^-1.Q-TOF/MS with electrospray ionization (ESI) and scanning range of m/z 50-1 200 were applied for analysis under positive and negative ion mode,respectively.All ionic peaks were assigned by comparison of reference substances,mass spectra data,database matching and literature reference,changes of chemical components in Salviae Miltiorrhizae Radix et Rhizoma were investigated by comparing number and area of ionic peaks before and after processing.Result: A total of 59 components were identified from raw products and characteristic processed products,and peak areas of 25 components showed obvious change.However,there were no new compound was found in characteristic processed products.After being processed with porcine cardiac blood,the contents of water-soluble ingredients (protocatechuic aldehyde,salvianolic acid C,F,G),fat-soluble ingredients (tanshinaldehyde,tanshindiol A,tanshinone Ⅰ) and amino acid (L-phenylalanine) in Salviae Miltiorrhizae Radix et Rhizoma were significantly increased.Conclusion: Changes of contents of chemical components in raw products and processed products of Salviae Miltiorrhizae Radix et Rhizoma are remarkable,part of water-soluble ingredients,fat-soluble ingredients and amino acids have quantitative change,which may be related to promoting treatment of Salviae Miltiorrhizae Radix et Rhizoma on cerebral ischemia after being processed with porcine cardiac blood.

BACKGROUND: Some porous materials have been developed to enhance biologic fusion of the implants to bone in spine fusion surgeries. However, there are several inherent limitations. In this study, a novel biomedical porous tantalum was applied to in vitro and in vivo experiments to test its biocompatibility and osteocompatibility. METHODS: Bone marrow-derived mesenchymal stem cells (BMSCs) were cultured on porous tantalum implant. Scanning electron microscope (SEM) and Cell Counting Kit-8 assay were used to evaluate the cell toxicity and biocompatibility. Twenty-four rabbits were performed discectomy only (control group), discectomy with autologous bone implanted (autograft group), and discectomy with porous tantalum implanted (tantalum group) at 3 levels: L3-L4, L4-L5, and L5-L6 in random order. All the 24 rabbits were randomly sacrificed at the different post-operative times (2, 4, 6, and 12 months; n = 6 at each time point). Histologic examination and micro-computed tomography scans were done to evaluate the fusion process. Comparison of fusion index scores between groups was analyzed using one-way analysis of variance. Other comparisons of numerical variables between groups were made by Student t test. RESULTS: All rabbits survived and recovered without any symptoms of nerve injury. Radiographic fusion index scores at 12 months post-operatively between autograft and tantalum groups showed no significant difference (2.89 ± 0.32 vs. 2.83 ± 0.38, F = 244.60, P = 0.709). Cell Counting Kit-8 assay showed no significant difference of absorbance values between the leaching liquor group and control group (1.25 ± 0.06 vs. 1.23 ± 0.04, t = -0.644, P = 0.545), which indicated the BMSC proliferation without toxicity. SEM images showed that these cells had irregular shapes with long spindles adhered to the surface of tantalum implant. No implant degradation, wear debris, or osteolysis was observed. Histologic results showed solid fusion in the porous tantalum and autologous bone implanted intervertebral spaces. CONCLUSION This novel porous tantalum implant showed a good biocompatibility and osteocompatibility, which could be a valid biomaterial for interbody fusion cages.
Animals ; Cell Proliferation ; physiology ; Diskectomy ; Lumbar Vertebrae ; surgery ; Microscopy, Electron, Scanning ; Prostheses and Implants ; Rabbits ; Spinal Fusion ; Tantalum ; chemistry

The aim of this paper was to explore the effects and possible mechanisms in vitro of tea polyphenols (TP) delaying human glomerular mesangial cells (HGMCs) senescence induced by high glucose (HG). HGMCs were cultured in vitro and divided into the normal group (N, 5.5 mmol·L⁻¹ glucose), the mannitol group(MNT, 5.5 mmol·L⁻¹ glucose plus 24.5 mmol·L⁻¹ mannitol), the high dose of D-glucose group (HG, 30 mmol·L⁻¹ glucose), the low dose of TP group (L-TP, 30 mmol·L⁻¹ glucose plus 5 mg·L⁻¹ TP) and the high dose of TP group (H-TP, 30 mmol·L⁻¹ glucose plus 20 mg·L⁻¹ TP), which were cultured in 5% CO₂ at 37 °C, respectively. Firstly, the effects of TP on the cell morphology of HGMCs were observed after 72 h-intervention. Secondly, the cell cycle, the positive rate of senescence-associated-β-galactosidase (SA-β-gal) staining and the telomere length were detected, respectively. Finally, the protein expressions of p53, p21 and Rb in the p53-p21-Rb signaling pathway were investigated, respectively. And the expressions of p-STAT3 and miR-126 were examined severally. The results indicated that HG not only arrested the cell cycle in G₁ phase but also increased the positive rate of SA-β-gal staining, and shortened the telomere length. HG led to the protein over-expressions of p53, p21 and Rb and HGMCs senescence by activating the p53-p21-Rb signaling pathway. In addition, L-TP delayed HGMCs senescence by improving the cell cycle G₁ arrest, reducing SA-β-gal staining positive rate and lengthening the telomere length. L-TP reduced the protein over-expressions of p53, P21 and Rb induced by HG and inhibited the telomere-p53-p21-Rb signaling pathway. Moreover, the expression of p-STAT3 was increased and the expression of miR-126 was decreased in HGMCs induced by HG. L-TP reduced the expression of p-STAT3 and increased the expression of miR-126 in HGMCs. In conclusion, HG could induce HGMCs senescence by activating the telomere-p53-p21-Rb signaling pathway in vitro. L-TP could delay HGMCs senescence through regulating STAT3/miR-126 expressions and inhibiting the telomere-p53-p21-Rb signaling pathway activation. These findings could provide the effective interventions in clinic for preventing and treating renal cell senescence in diabetic kidney disease.
Cells, Cultured ; Cellular Senescence ; Cyclin-Dependent Kinase Inhibitor p21 ; Glucose ; Humans ; Mesangial Cells ; MicroRNAs ; Polyphenols ; STAT3 Transcription Factor ; Tea ; Telomere ; Tumor Suppressor Protein p53

OBJECTIVE Lychee seed, a famous traditional Chinese medicine, recently were reported to improve the learning and memory abilities in mice. However, it is still unclear whether lychee seed saponins (LSS) can improve the cognitive function and associated mechanisms. METHODS In present studies, we established the Alzheimer disease (AD) model by injecting Aβ25-35 into the lateral ventricle of rats. Then the spatial learning and memory abilities of LSS- treated rats were evaluated with the Morris water maze, meanwhile the protein expressions of AKT, GSK3β and Tau in the hippo?campal neuron were analyzed by immunohistochemistry and Western blotting. RESULTS The results showed LSS can improve the cognitive functions of AD rats through shortening the escape latency, increasing the number across the platform, platform quadrant dwell time and the percentage of the total distance run platform quadrant. The protein expression of AKT was significantly up-regulated and that of GSK3β and Tau were decreased remarkably in the hippocampal CA1 area. CONCLUSION Our study is the first to show that LSS significantly improve the cognitive function and prevent hippocampal neuronal injury of the rats with AD by activation of the PI3K/AKT/GSK3β signaling pathway, suggesting LSS may be developed into the nutrient supplement for the treatment of AD.

Objective To compare characteristics of stress variations in 3D finite element models of normal and degenerative lumbar vertebrae and the dose-effect relationship, and analyze the mechanism of mechanical balance by traditional Chinese medicine (TCM) manipulation on degenerative lumbar vertebrae. Methods The 3D finite element model of intact, real human degenerative lumbar vertebrae (L4-5) was established to simulate the physiological activity of flexion and extension in lumbar vertebrae. The characteristics of stress variation in degenerative lumbar vertebrae under external loading, namely, the TCM manipulation was analyzed, and the stress variation in degenerative lumbar vertebrae under gradual increasing-external loading was analyzed as well, which was compared with the stress and strain variation in normal lumbar vertebrae under different motion status. Results Under different motion status, the stress distributions on lumbar disc as well as the elastic modulus of nucleus pulposus and fiber ring showed a gradually increasing tendency with lumbar degeneration increasing. TCM manipulation could change the stress distributions on lumbar disc, enlarge the space of spinal canal to a certain degree, and decrease the stress on nerve root. Stresses on small joints of the vertebral body and vertebral pedicle under posterior extension were larger than those under anterior flexion, while stresses on intervertebral disc under anterior flexion were greater than those under posterior extension, which showed a gradually increasing trend from top to bottom. Conclusions The mechanical environment of human lumbar vertebrae can be balanced by TCM manipulation, for the purpose of improving and treating lumbar disc diseases. The comparison with the 3D finite element model of normal human lumbar vertebrae and investigation on lumbar degeneration from perspective of changes in biomechanical environment and characteristics can provide scientific basis for clinic application of TCM manipulation in prevention and treatment of lumbar degenerative diseases, as well as new research idea for studying mechanical mechanism of TCM manipulation in effective prevention and treatment of lumbar lesions.

OBJECTIVETo investigate the safety and effectiveness of autologous hematopoietic stem cell transplantation (auto-HSCT) using tumor-ablative conditioning regiment for patients with refractory/relapsed non-Hodgkin's lymphoma.

METHODSThe clinical data of 16 patients with refractory/relapsed non-Hodgkin's lymphoma received above-mentioned therapeutic regimen from January 2013 to July 2015 was analyzed retrospectively, and conditioning-related toxicity, engraftment, infection, relapse and survival rate were evaluated.

RESULTSNo conditioning-related organs' failure and mortality were found. Only 1 patient had not been engrafted, and the engraftment rate was 93.7%. The incidence of serious infection was 31.2%. The median follow-up was 20.5(1-30) months, and 3 patients died, out of them 2 patients died of relapse. Two year overall survival (OS) , disease-free survival (DFS) and relapse rates were 80.2%, 74.5% and 20.6% respectively.

CONCLUSIONAuto-HSCT using tumor-ablative conditioning regimen is safe and effective for patients with refractory/relapsed non-Hodgkin's lymphoma, and it possess a certain effect for reducing disease relapse after transplantation.