Nattokinase's Neuroprotective Mechanisms in Ischemic Stroke: Targeting Inflammation, Oxidative Stress, and Coagulation.

Aims: Nattokinase (NK), a potent serine endopeptidase, has exhibited a variety of pharmacological effects, including thrombolysis, anti-inflammation, and antioxidative stress. Building on previous research highlighting NK's promise in nerve regeneration, our study investigated whether NK exerted protective effects in transient middle cerebral artery occlusion (tMCAO)-induced cerebral ischemia-reperfusion injury and the underlying mechanisms. Results: The rats were administered NK (5000, 10000, 20000 FU/kg, i.g., 7 days before surgery, once daily). We showed that NK treatment dose dependently reduced the infarction volume and improved neurological symptoms, decreased the proinflammatory and coagulation cytokines levels, and attenuated reactive oxygen species (ROS) in the infarcted area of tMCAO rats. We also found that NK could exert neuroprotective effects in a variety of vitro models, including the microglia inflammation model and neuronal oxygen-glucose deprivation/reperfusion (OGD/R) model. Notably, NK effectively countered OGD/R-induced neuron death, modulating diverse pathways, including autophagy, apoptosis, PARP-dependent death, and endoplasmic reticulum stress. Furthermore, the neuroprotection of NK was blocked by phenylmethylsulfonyl fluoride (PMSF), a serine endopeptidase inhibitor. We revealed that heat-inactive NK was unable to protect against tMCAO injury and other vitro models, suggesting NK attenuated ischemic injury by its enzymatic activity. We conducted a proteomic analysis and found inflammation and coagulation were involved in the occurrence of tMCAO model and in the therapeutic effect of NK. Innovation and Conclusion: In conclusion, these data demonstrated that NK had multifaceted neuroprotection in ischemic brain injury, and the therapeutic effect of NK was related with serine endopeptidase activity.

Early diagnosis of renal pelvis villous adenoma: A case report.

BACKGROUND: Villous adenoma is a rare tumor in the urinary system that usually occurs in the bladder. It is extremely uncommon in the renal pelvis. Most of the previously reported cases have been diagnosed with severe hydronephrosis associated with renal parenchyma atrophy prior to surgery. Because of its rarity, available information on the pathogenesis, diagnosis, treatment and prognosis of the disease is limited. We reported a case of kidney stones with hydronephrosis. During percutaneous nephroscopic lithotripsy, a renal pelvis tumor was found. Biopsy confirmed that the tumor was a villous adenoma of the renal pelvis. CASE SUMMARY: A 68-year-old female was admitted to the hospital due to right kidney stones with right hydronephrosis. After admission, a urinary system plain computed tomography scan was performed, which revealed right kidney stones with right hydronephrosis and right upper ureteral dilatation. Multiple new cauliflower-like papillary masses were then discovered in the renal pelvis and calyces during right percutaneous nephroscopic lithotripsy. Biopsy results indicated villous adenoma with high-grade glandular intraepithelial neoplasia. The patient underwent laparoscopic radical resection of the right kidney and ureter. Based on histopathological and immunohistochemical examination, the patient was diagnosed with villous adenoma without adenocarcinoma. CONCLUSION: Villous adenoma is rare in the urinary system. We reported a case of renal pelvis villous adenoma, which may provide useful information for the early diagnosis and treatment of this tumor.

Inhibition of ASGR1 decreases lipid levels by promoting cholesterol excretion.

High cholesterol is a major risk factor for cardiovascular disease1. Currently, no drug lowers cholesterol through directly promoting cholesterol excretion. Human genetic studies have identified that the loss-of-function Asialoglycoprotein receptor 1 (ASGR1) variants associate with low cholesterol and a reduced risk of cardiovascular disease2. ASGR1 is exclusively expressed in liver and mediates internalization and lysosomal degradation of blood asialoglycoproteins3. The mechanism by which ASGR1 affects cholesterol metabolism is unknown. Here, we find that Asgr1 deficiency decreases lipid levels in serum and liver by stabilizing LXRα. LXRα upregulates ABCA1 and ABCG5/G8, which promotes cholesterol transport to high-density lipoprotein and excretion to bile and faeces4, respectively. ASGR1 deficiency blocks endocytosis and lysosomal degradation of glycoproteins, reduces amino-acid levels in lysosomes, and thereby inhibits mTORC1 and activates AMPK. On one hand, AMPK increases LXRα by decreasing its ubiquitin ligases BRCA1/BARD1. On the other hand, AMPK suppresses SREBP1 that controls lipogenesis. Anti-ASGR1 neutralizing antibody lowers lipid levels by increasing cholesterol excretion, and shows synergistic beneficial effects with atorvastatin or ezetimibe, two widely used hypocholesterolaemic drugs. In summary, this study demonstrates that targeting ASGR1 upregulates LXRα, ABCA1 and ABCG5/G8, inhibits SREBP1 and lipogenesis, and therefore promotes cholesterol excretion and decreases lipid levels.

The Effect of Calycosin-7-O-ß-D-Glucoside and its Synergistic Augmentation of Cisplatin-induced Apoptosis in SK-OV-3 Cells.

OBJECTIVE: This study aims to examine the synergetic augmentation of calycosin-7-O-ß-D-glucoside (CG) on cisplatin (CDDP) to induce apoptosis of human epithelial ovarian SK-OV-3 cancer cells. METHODS: The SK-OV-3 cells were divided into four groups: control, CDDP monotherapy, CG monotherapy, and combined CDDP and CG treatment. The cell counting kit-8 method detected cell proliferation at different times and under different treatments. Hoechst 33258 staining and annexin V-FITC/propidium iodide double staining methods were used to observe the apoptosis of the SK-OV-3 cells. The caspase-3 enzyme activity detection method, quantitative reverse transcription-polymerase chain reaction, and western blot were used to detect the apoptosis-related factors and the activities of the enzyme in SK-OV-3 cells. RESULTS: The inhibition rates of SK-OV-3 cell proliferation when exposed to 10 µM of CDDP, 50 µM of CG, and a combination of 10 µM of CDDP and 50 µM of CG were 23.2% ± 1.1%, 26.7% ± 2.0%, and 46.7% ± 1.3% after 48 h, respectively. Following the use of the drug combination, the apoptosis rate and caspase-3 enzyme activity were significantly higher than in the single-drug treatment group; the data differences were also significant (p < 0.05). At the protein and ribonucleic acid levels, CG significantly enhanced the effect of CDDP on p53, caspase-3, caspase-9, Bax, and Bcl-2. CONCLUSION: In vitro, CG significantly increases the CDDP-induced apoptosis of the SK-OV-3 cells through the p53 pathway at the cellular level. In addition, using the drugs in combination reduces the toxicity and side effects caused by using CDDP alone.

Comparison of serum inflammatory indicators and radiographic results in MAKO robotic-assisted versus conventional total knee arthroplasty for knee osteoarthritis: a retrospective study of Chinese patients.

BACKGROUND: The purpose of this study was to compare the serum inflammatory indicators and radiographic results of conventional manual total knee arthroplasty (CM-TKA) with those of MAKO-robotic assisted total knee arthroplasty (MA-TKA). METHODS: We retrospectively analysed 65 patients with knee osteoarthritis who underwent unilateral TKA from December 2020 to November 2021 in our department, which included 34 patients who underwent MA-TKA and 31 patients who underwent CM-TKA. The tourniquet time and estimated blood loss (EBL) were compared between the two groups. Knee function was evaluated using range of motion (ROM), functional score and pain score. Leukocytes, C-reactive protein (CRP), erythrocyte sedimentation rate (ESR), interleukin-6 (IL-6), creatine kinase (CK), and neutrophil-to-lymphocyte ratio (NLR) were recorded at 3 time points (preoperative, and on the first and third postoperative days). The hip-knee-ankle angle (HKA) and the femoral and tibial component angles in the coronal and sagittal planes were used for postoperative radiographic evaluation. RESULTS: The postoperative MA-TKA group had less EBL (496.9 ± 257.8 vs. 773.0 ± 301.3 ml, p < 0.001). There was no significant difference in knee function scores at 6 weeks postoperatively (p > 0.05). IL-6 levels were significantly lower in the MA-TKA group on the 1st postoperative day (11.4 (5.2, 21.0) vs. 24.6 (86.3, 170.8), p = 0.031). This difference in inflammatory indices became more pronounced at 72 hours after the operation because CRP, ESR, IL-6, and CK values were significantly lower in the MA-TKA group on the 3rd postoperative day (72 h) (p < 0.05). Postoperative radiographic examinations performed 2 days after the MA-TKA group suggested that only 2 cases of HKA had outlier values, which was remarkably better than the 12 cases found in the CM-TKA group (5.9% vs. 38.7%, p < 0.001). The frontal femoral component was significantly closer to the expected value of 90° in the MA-TKA group (90.9 (90.5, 92.3) vs. 92.4 (91.3, 93.7), p = 0.031). The remaining imaging evaluation parameters were not significantly different between the two groups (p > 0.05). CONCLUSIONS: In Chinese patients with OA, there was a milder systemic inflammatory response in the early postoperative period after MA-TKA compared to that of CM-TKA, as well as better radiographic outcomes. However, the tourniquet time was prolonged, and no advantages were observed in terms of functional score or pain score in the short-term follow-up.

Gender Differences in Ankylosing Spondylitis Patients with Advanced Hip Involvement: Results from A Matched Retrospective Cohort Study.

OBJECTIVE: To determine the gender differences in ankylosing spondylitis (AS) patients with advanced hip involvement. METHODS: We retrospectively analyzed the 373 consecutive AS patients with advanced hip involvement from 2012 to 2017 and divided them into two groups by sex with 340 men and 33 women. Research data on hip involvement in the patients were obtained from medical records and radiographs. The severity of radiographic hip involvement was evaluated by the Bath Ankylosing Spondylitis Radiology Hip Index (BASRI-hip) scoring system. The data on clinical characteristics, radiographic hip involvement, and laboratory values were compared between the two groups. The comparison was performed again between the two groups after adjusting for the onset age and disease duration by propensity score matching (PSM). RESULTS: Men underwent total hip arthroplasty earlier than women in the patients, with a median age of 31 years (range, 19-67 years) vs 36 years (range, 23-67 years), respectively (P < 0.05). Hip involvement was found to be younger in men than that in women, with a median age of 18 years (range, 7-56 years) vs 23 years (range, 5-55 years) (P < 0.05), and men with bilateral onset in hips had a higher frequency than women with that (66.2% vs 39.4%) (P < 0.05). There was no gender difference in the proportion of bilateral advanced hip involvement (85.3% vs 72.7%) (P > 0.05). The proportion of the patients who had spinal involvement (89.1% vs 69.7%), flexion contracture in the hip (43.8% vs 24.2%), hip range of motion =0° (53.5% vs 30.3%), and an elevated level of C-reactive protein (CRP) (69.1% vs 51.5%) was significantly higher in men than that in women (P < 0.05). After adjusting for the onset age and disease duration by PSM (1:1), men with bilateral onset in hips still had a higher frequency than women with that (76.7% vs 40.0%), and the proportion of the patients who had spinal involvement (90.0% vs 66.7%) and an elevated level of CRP (80.0% vs 53.3%) was significantly higher in men than that in women (P < 0.05). CONCLUSIONS: The disease pattern of hip involvement in AS has gender differences, with bilateral onset being the dominant pattern in men and unilateral onset being more common in women. However, the frequency of bilateral advanced hip involvement has no gender difference eventually. The higher prevalence of spinal involvement in men with AS may be responsible for the more severe functional impairment compared with women.

SUMOylation of the ubiquitin ligase IDOL decreases LDL receptor levels and is reversed by SENP1.

Inducible degrader of the low-density lipoprotein receptor (IDOL) is an E3 ubiquitin ligase mediating degradation of low-density lipoprotein (LDL) receptor (LDLR). IDOL also controls its own stability through autoubiquitination, primarily at lysine 293. Whether IDOL may undergo other forms of posttranslational modification is unknown. In this study, we show that IDOL can be modified by small ubiquitin-like modifier 1 at the K293 residue at least. The SUMOylation of IDOL counteracts its ubiquitination and augments IDOL protein levels. SUMOylation and the associated increase of IDOL protein are effectively reversed by SUMO-specific peptidase 1 (SENP1) in an activity-dependent manner. We further demonstrate that SENP1 affects LDLR protein levels by modulating IDOL. Overexpression of SENP1 increases LDLR protein levels and enhances LDL uptake in cultured cells. On the contrary, loss of SENP1 lowers LDLR levels in an IDOL-dependent manner and reduces LDL endocytosis. Collectively, our results reveal SUMOylation as a new regulatory posttranslational modification of IDOL and suggest that SENP1 positively regulates the LDLR pathway via deSUMOylation of IDOL and may therefore be exploited for the treatment of cardiovascular disease.

[Value of Red Blood Cell Distribution Width and Fibrinogen Level for Evaluation of the Therapeutic Efficacy and Prognosis in Patients with Diffuse Large B-Cell Lymphoma].

OBJECTIVE: To investigate the value of red blood cell distribution width (RDW) and fibrinogen (Fib) level for the evaluation of therapeutic efficacy and prognosis in patients with diffuse large B-cell lymphoma (DLBCL). METHODS: The relationship between RDW/Fib at initial diagnosis and efficacy and the clinical outcome was retro-spectively analyzed based on the study of 105 patients with DLBCL. The patients were divided into two groups: low RDW group (≤15%) and high RDW group (>15%), low Fib group (Fib≤4 g/L) and high Fib group (Fib>4 g/L) according to the normal values of RDW and Fib. Therapeutic efficacy, overall survival (OS) time and progression free survival (PFS) time were compared between two groups. The correlation between each factors and efficacy, prognosis was analyzed by univariate and multivariate regression. RESULTS: The therapeutic efficacy (P<0.001), OS time(P=0.004), and PFS time(P=0.007) were poorer in the high RDW group as compared with the low RDW group. The efficacy (P=0.015) and PFS time(P=0.04) were poorer in the high Fib group as compared with the low Fib group. Multivariate analysis showed that high RDW was the independent risk factor for efficacy of DLBCL patients (OR=3.394, 95% CI 1.093-10.539, P=0.035). CONCLUSION: High RDW and high Fib associate with poor efficacy in DLBCL patients.

Lithium Chloride-Releasing 3D Printed Scaffold for Enhanced Cartilage Regeneration.

BACKGROUND We synthetized a 3D printed poly-ε-caprolactone (PCL) scaffold with polydopamine (PDA) coating and lithium chloride (LiCl) deposition for cartilage tissue engineering and analyzed its effect on promoting rabbit bone marrow mesenchymal stem cells (rBMSC) chondrogenesis in vitro. MATERIAL AND METHODS PCL scaffolds were prepared by 3D printing with a well-designed CAD digital model, then modified by PDA coating to produce PCL-PDA scaffolds. Finally, LiCl was deposited on the PDA coating to produce PCL-PDA-Li scaffolds. The physicochemical properties, bioactivity, and biocompatibility of PCL-PDA-Li scaffolds were accessed by comparing them with PCL scaffolds and PCL-PDA scaffolds. RESULTS 3D PCL scaffolds exhibited excellent mechanical integrity as designed. PDA coating and LiCl deposition improved surface hydrophilicity without sacrificing mechanical strength. Li⁺ release was durable and ion concentration did not reach the cytotoxicity level. This in vitro study showed that, compared to PCL scaffolds, PCL-PDA and PCL-PDA-Li scaffolds significantly increased glycosaminoglycan (GAG) formation and chondrogenic marker gene expression, while PCL-PDA-Li scaffolds showed far higher rBMSC viability and chondrogenesis. CONCLUSIONS 3D printed PCL-PDA-Li scaffolds promoted chondrogenesis in vitro and may provide a good method for lithium administration and be a potential candidate for cartilage tissue engineering.

TGF-ß1-PML SUMOylation-peptidyl-prolyl cis-trans isomerase NIMA-interacting 1 (Pin1) form a positive feedback loop to regulate cardiac fibrosis.

Transforming growth factor-ß (TGF-ß) signaling pathway is involved in fibrosis in most, if not all forms of cardiac diseases. Here, we evaluate a positive feedback signaling the loop of TGF-ß1/promyelocytic leukemia (PML) SUMOylation/Pin1 promoting the cardiac fibrosis. To test this hypothesis, the mice underwent transverse aortic constriction (3 weeks) were developed and the morphological evidence showed obvious interstitial fibrosis with TGF-ß1, Pin1 upregulation, and increase in PML SUMOylation. In neonatal mouse cardiac fibroblasts (NMCFs), we found that exogenous TGF-ß1 induced the upregulation of TGF-ß1 itself in a time- and dose-dependent manner, and also triggered the PML SUMOylation and the formation of PML nuclear bodies (PML-NBs), and consequently recruited Pin1 into nuclear to colocalize with PML. Pharmacological inhibition of TGF-ß signal or Pin1 with LY364947 (3 µM) or Juglone (3 µM), the TGF-ß1-induced PML SUMOylation was reduced significantly with downregulation of the messenger RNA and protein for TGF-ß1 and Pin1. To verify the cellular function of PML by means of gain- or loss-of-function, the positive feedback signaling loop was enhanced or declined, meanwhile, TGF-ß-Smad signaling pathway was activated or weakened, respectively. In summary, we uncovered a novel reciprocal loop of TGF-ß1/PML SUMOylation/Pin1 leading to myocardial fibrosis.

PMLIV overexpression promotes TGF-ß-associated epithelial-mesenchymal transition and migration in MCF-7 cancer cells.

The epithelial-mesenchymal transition (EMT) is a key event associated with metastasis and dissemination in breast tumor pathogenesis. Promyelocytic leukemia (PML) gene produces several isoforms due to alternative splicing; however, the biological function of each specific isoform has yet to be identified. In this study, we report a previously unknown role for PMLIV, the most intensely studied nuclear isoform, in transforming growth factor-ß (TGF-ß) signaling-associated EMT and migration in breast cancer. This study demonstrates that PMLIV overexpression promotes a more aggressive mesenchymal phenotype and increases the migration of MCF-7 cancer cells. This event is associated with activation of the TGF-ß canonical signaling pathway through the induction of Smad2/3 phosphorylation and the translocation of phospho-Smad2/3 to the nucleus. In this study, we report a previously unknown role for PMLIV in TGF-ß signaling-induced regulation of breast cancer-associated EMT and migration. Targeting this pathway may be therapeutically beneficial.

Circulating microRNAs as novel biomarkers of ALK-positive nonsmall cell lung cancer and predictors of response to crizotinib therapy.

Circulating microRNAs are potential diagnostic and predictive biomarkers, but have not been investigated for patients with anaplastic lymphoma kinase (ALK)-positive lung cancer. In this exploratory study, we sought to identify potential plasma biomarkers for ALK-positive non-small cell lung cancer (NSCLC). A microRNA microarray was used to select ALK-related microRNAs in ALK-positive NSCLC (n = 3), ALK-negative NSCLC (n = 3), and healthy subjects (n = 3). Plasma levels of 21 microRNAs were differentially expressed for ALK-positive and ALK-negative NSCLC, including 14 down-regulated and 7 up-regulated microRNAs. We also identified 5s rRNA as the most stable endogenous control gene using geNorm and NormFinder algorithms. Candidate microRNAs in plasma from ALK-positive (n = 41) and ALK-negative NSCLC patients (n = 32) were quantified using real-time reverse transcriptase quantitative polymerase chain reaction. The expression levels of miR-28-5p, miR-362-5p, and miR-660-5p were all down-regulated in ALK-positive NSCLC, compared with ALK-negative NSCLC. The areas under the receiver operating characteristic curves of miR-28-5p, miR-362-5p, miR-660-5p, and 3-microRNAs panel were 0.873, 0.673, 0.760, and 0.876, respectively. The positive predictive values of miR-28-5p, miR-362-5p, and miR-660-5p were 96.43%, 80.77%, and 83.87%, respectively. Increased plasma levels of miR-660-5p after crizotinib treatment predicted good tumor response (p = 0.012). The pre-crizotinib levels of miR-362-5p were significantly associated with progression-free survival (p = 0.015). Thus, in this preliminary investigation, we identified a potential panel of 3 microRNAs for distinguishing between patients with ALK-positive and ALK-negative NSCLC. We also identified miR-660-5p and miR-362-5p as potential predictors for response to crizotinib treatment.

Preparation of gentamicin-loaded electrospun coating on titanium implants and a study of their properties in vitro.

PURPOSE: Implant-related infections are disastrous complications in the clinic, and there are no effective therapies. In this preliminary study, gentamicin-loaded coating on titanium implants was prepared using the electrospinning technique, and some properties of the coating titanium implants were studied. METHODS: We adopted the electrospinning technique to prepare gentamicin-coated titanium implants. The surface structure of the coating implants was observed using scanning electron microscope. An elution study was performed to determine the release behavior of the gentamicin from the coating. The antibacterial efficacy and quantitative analysis of the bacterial adhesion of Staphylococcus aureus were evaluated in vitro, and the cytotoxicity of the coated titanium implants on osteoblasts was investigated in vitro. RESULTS: The morphology of the gentamicin-coated titanium implants exhibited nanofibers, and the release of gentamicin showed an initial gentamicin burst followed by a slow release. The gentamicin-coated titanium implants had a persistent antibacterial efficacy for 1 week and significantly reduced the adhesion of the Staphylococcus aureus compared with bare titanium implants in vitro. There was no cytotoxicity observed in vitro for the gentamicin-coated implants. CONCLUSION: The gentamicin-coated titanium implants, which were prepared using an electrospinning technique, present many advantages and may be considered to prevent and treat the implant-related infections.

Radiotherapy of unicentric mediastinal Castleman's disease.

Castleman's disease is a slowly progressive and rare lymphoproliferative disorder. Here, we report a 55-year-old woman with superior mediastinal Castleman's disease being misdiagnosed for a long term. We found a 4.3 cm mass localized in the superior mediastinum accompanied with severe clinical symptoms. The patient underwent an exploratory laparotomy, but the mass failed to be totally excised. Pathologic examination revealed a mediastinal mass of Castleman's disease. After radiotherapy of 30 Gy by 15 fractions, the patient no longer presented previous symptoms. At 3 months after radiotherapy of 60 Gy by 30 fractions, Computed tomography of the chest showed significantly smaller mass, indicating partial remission. Upon a 10-month follow-up, the patient was alive and free of symptoms.

[Clinical significance of expressions of tumor markers in peripheral blood in non-small cell lung cancer].

OBJECTIVE: To investigate the expressions of BJ-TSA-9, CK19 and Pre-proGRP mRNA in peripheral blood from the patients with non-small cell lung cancer and analyze their correlations with non-small cell lung cancer. METHODS: The expressions of BJ-TSA-9, CK19 and Pre-proGRP mRNA were detected by nested reverse transcription-PCR assay in peripheral blood from the patients with non-small cell lung cancer (n = 120), benign pulmonary disease (n = 106) and from healthy subjects (n = 80) so as to further investigate their relationship with clinicopathological features and prognosis. Meantime we also examined the sensitivity, specificity and accuracy of combination detection. RESULTS: The expressions of BJ-TSA-9, CK19 and Pre-proGRP mRNA in non-small cell lung cancer patients were 56.7%, 57.5%, 35.0%, higher than that of benign pulmonary disease (0.9%, 6.6%, 5.7%) and healthy groups (0, 3.8%, 0, all P < 0.05). The ROC curves indicated the sensitivity of combined detection was 84.3% and the specificity of combined detection was 94.6%. Univariate analysis revealed that the clinical stage, the ECOG score and the number of positive marker had significant association with overall survival (OS) (χ(2) = 67.928, 95.981, 60.285, all P = 0.000). Multivariate analysis indicated that the clinical stage, ECOG score and the number of positive marker was an independent prognostic factor each (HR = 2.866, 4.251, 1.845, all P = 0.000). CONCLUSION: BJ-TSA-9, CK19 and Pre-proGRP mRNA may be the specific and sensitive markers to detect circulating tumor cells in the peripheral blood of non-small cell lung cancer patients.