GroEL triggers NLRP3 inflammasome activation through the TLR/NF-κB p-p65 axis in human periodontal ligament stem cells.

The interaction between bacteria and the host plays a vital role in the initiation and progression of systemic diseases, including gastrointestinal and oral diseases, due to the secretion of various virulence factors from these pathogens. GroEL, a potent virulence factor secreted by multiple oral pathogenic bacteria, is implicated in the damage of gingival epithelium, periodontal ligament, alveolar bone and other peripheral tissues. However, the underlying biomechanism is still largely unknown. In the present study, we verify that GroEL can trigger the activation of NLRP3 inflammasome and its downstream effector molecules, IL-1ß and IL-18, in human periodontal ligament stem cells (hPDLSCs) and resultantly induce high activation of gelatinases (MMP-2 and MMP-9) to promote the degradation of extracellular matrix (ECM). GroEL-mediated activation of the NLRP3 inflammasome requires the participation of Toll-like receptors (TLR2 and TLR4). High upregulation of TLR2 and TLR4 induces the enhancement of NF-κB (p-p65) signaling and promotes its nuclear accumulation, thus activating the NLRP3 inflammasome. These results are verified in a rat model with direct injection of GroEL. Collectively, this study provides insight into the role of virulence factors in bacteria-induced host immune response and may also provide a new clue for the prevention of periodontitis.

Development of a deep learning-based model to diagnose mixed-type gastric cancer accurately.

OBJECTIVE: The accurate diagnosis of mixed-type gastric cancer from pathology images presents a formidable challenge for pathologists, given its intricate features and resemblance to other subtypes of gastric cancer. Artificial Intelligence has the potential to overcome this hurdle. This study aimed to leverage deep machine learning techniques to establish a precise and efficient diagnostic approach for this cancer type which can also predict the metastatic risk using two software, U-Net and QuPath, which have not been trialled in gastric cancers. METHODS: A U-Net neural network was trained to recognise, and segment differentiated components from 186 pathology images of mixed-type gastric cancer. Undifferentiated components in the same images were annotated using the open-source pathology imaging software QuPath. The outcomes from U-Net and QuPath were used to calculate the ratios of differentiation/undifferentiated components which were correlated to lymph node metastasis. RESULTS: The models established by U-Net recognised ∼91% of the regions of interest, with precision, recall, and F1 values of 90.2%, 90.9% and 94.6%, respectively, indicating a high level of accuracy and reliability. Furthermore, the receiver operating characteristic curve analysis showed an area under the cure of 91%, indicating good performance. A bell-curve correlation between the differentiated/undifferentiated ratio and lymphatic metastasis was found (highest risk between 0.683 and 1.03), which is paradigm-shifting. CONCLUSION: U-Net and QuPath exhibit promising accuracy in the identification of differentiated and undifferentiated components in mixed-type gastric cancer, as well as paradigm-shifting prediction of metastasis. These findings bring us one step closer to their potential clinical application.

Supraorbital eyebrow keyhole approach for microsurgical management of ruptured anterior communicating artery aneurysm.

The mortality and disability rate of patients with ruptured anterior communicating artery (AComA) aneurysm after bleeding is high. Even with the most advanced treatment methods, the incidence of complications remains high. The purpose of the present study was to determine the efficacy of microsurgery via supraorbital eyebrow keyhole approach (SOEK) in clipping ruptured AComA aneurysms. Between September 2010 and October 2018, 543 patients with intracranial aneurysms were admitted to the Department of Neurosurgery of the Third Affiliated Hospital of Sun Yat-Sen University (Guangzhou, China). Among them, 85 patients with ruptured AComA aneurysm and subarachnoid hemorrhage (SAH) underwent microsurgical clipping via the SOEK approach. In those patients, the clipping rate, complications and clinical efficacy of treatment were evaluated. The average age of the patients was 52.69±9.94 years (range, 28-78 years). The proportions of small, medium and large aneurysms were 83.5, 15.3 and 1.2%, respectively. Procedural complications occurred in 9 cases (10.5%). The occlusion rate of the aneurysms was 98.8%. The average follow-up period was 37.9 (±24.5) months. A total of 81.2% of the patients with SAH had a good clinical prognosisat 1 year (modified Rankin scale score, ≤2). In conclusion, for a skilled and experienced surgeon, SOEK was indicated to be a safe procedure for the treatment of ruptured AComA aneurysms; it provided sufficient intra-operative exposure and a high clipping rate.

Molecular mechanism of microRNA-21 promoting Schwann cell proliferation and axon regeneration during injured nerve repair.

At present, the functional recovery after nerve injury is not satisfactory in clinical practice. The aim of this study was to explore the molecular mechanism of miR-21 promoting Schwann cells (SC) proliferation and axon regeneration after peripheral nerve injury, providing a theoretical basis for injured nerve repair. Nerve injury models were constructed to determine the expression of miR-21 in the injured nerve by Quantitative Real-Time PCR (qRT-PCR). After miR-21 over-expression SC (mimic-miR-21) group, control SC (control-miR-21) group and blank SC (RSC96) group were constructed, SC proliferation was determined by CCK-8, cell cycle was analysed by flow cytometry, dorsal root ganglion neuron (DRGn) axon regeneration was observed after DRGn was cultured with SCs for 7 days, the expressions of TGFßI, TIMP3, EPHA4 as well as apoptosis-related proteins caspase-3 and caspase-9 were detected by qRT-PCR and Western blot in the three groups, respectively. Target genes were confirmed by dual-luciferase reporter gene assay. The expressions of TGFßI, TIMP3 and EPHA4 were assessed by immunofluorescence in vivo. qRT-PCR indicated that miR-21 expression was significantly higher in the model group than in the sham operation and blank groups. SC proliferation index (PI) was significantly higher, the apoptosis rate was significantly lower, the axon was significantly longer, and mRNA and protein expressions of TGFßI, TIMP3, EPHA4 as well as apoptosis-related proteins caspase-3 and caspase-9 were significantly lower in the mimic-miR-21 group than in the control-miR-21 and RSC96 groups. The double luciferase assay confirmed that TGFßI, TIMP3 and EPHA4 were potential target genes of miR-21. In vivo immunofluorescence also indicated that expressions of TGFßI, TIMP3, EPHA4 were lower in the mimic-miR-21 group than in the control-miR-21 and RSC96 groups. We conclude that during injured peripheral nerve repair, miRNA-21 plays an important role in promoting SC proliferation and axon regeneration by regulating TGFßI, TIMP3 and EPHA4 target genes.

miR-149 rs2292832 C allele enhances the cytotoxic effect of temozolomide against glioma cells.

Glioma is a common cancer that affects people worldwide with high morbidity and mortality. Human miR-149 rs2292832 C/T polymorphism and miR-149-5p expressions have been documented to play important roles in various type of cancers. This study aims to assess the impact of miR-149 rs2292832 C/T polymorphism and miR-149-5p expressions in cytotoxic effect of temozolomide against glioma cells. A total of 137 cases of glioma patients and 21 healthy cases were enrolled in this study for clinical research. We found that miR-149-5p was significantly downregulated in glioma cell lines and in blood leukocyte of glioma patients. Furthermore, miR-149 rs2292832 C/T polymorphism was significantly associated with glioma prognosis and temozolomide resistance. Subsequently, the glioma cell lines stable transfected with common miR-149 expression construct (miR-149-T) and the variant miR-149 expression construct (miR-149-C) were used to determine the regulatory effect of miR-149 rs2292832 C on glioma cells progression. Data revealed that miR-149 rs2292832 C allele could enhance the miR-149-5p expressions, and therefore, prevent the proliferation of glioma cells and increase the cytotoxicity of temozolomide against glioma cells. These functions of miR-149-C were demonstrated to be triggered by CDK6/SOX2 pathway inhibition. The above results demonstrated that miR-149 rs2292832 C/T polymorphism was a potential prognostic biomarker for glioma development by regulating miR-149/CDK6 axis.

Surgical treatment of foreign body embolus in the Middle cerebral artery secondary to neck injury.

Background: We report a case of a foreign body embolus to the middle cerebral artery and reviewed similar cases previously reported. Methods: A 30-year-old man was seen 72 days after a penetrating neck injury with a 1-month history of numbness in the left limb and impairment of the fine movement in the left hand. Radiological examination revealed a foreign body in the M2 portion of the right middle cerebral artery (MCA). The patient received arteriotomy and in situ suturing. Results: During the operation, we found a metallic foreign body at the bifurcation of the M2 upper trunk of the right MCA, narrowed distal blood vessels and thinned vessel walls. The foreign body was surrounded by granulation tissue. Both foreign body and granulation tissue were removed slowly followed by in situ suturing. Indocyanine green angiography confirmed arterial patency. Three days after the surgery, the patient developed numbness and weakness in the left arm, with a muscle strength of grade 4. Computed tomography showed partial infarction in the right temporal lobe. Then, antispasmodic drugs were used. Muscle strength recovered by 14 days after the operation. Conclusions: In the subacute stage, surgery can be conducted to remove intra-arterial foreign bodies along with their surrounding granulation tissue if computed tomography perfusion suggests a decreased blood flow reserve capacity.

Selenium nanoparticles reduce glucose metabolism and promote apoptosis of glioma cells through reactive oxygen species-dependent manner.

Gliomas are the most common, malignant, and lethal tumors in adults. Furthermore, gliomas are highly resistant to current chemotherapeutic drugs. Thus, new effective anticancer drugs for glioma are urgently needed. Selenium nanoparticles have been reported to have potent anti-tumor activity, although the specific mechanism is not fully understood. This study aimed to test the anti-tumor effect of selenium nanoparticles and its mechanism. We used selenium nanoparticles to treat commercial glioma cell lines, and patient-derived glioma cells, and then used the MTT assay to determine selenium nanoparticles effect against these. Apoptotic cell death was determined by annexin V-Fluos staining kit. Glucose uptake, lactate, and adenosine triphosphate production, together with hexokinase 2 and pyruvate kinase activities were measured to determine the glucose metabolism level. Reactive oxygen species production was tested using 2',7'-dichlorodihydrofluorescein diacetate. Our results showed that selenium nanoparticles had a potent cytotoxic effect in glioma cells, regardless of whether they were drug-resistant or not, whereas it showed less toxic effect in normal healthy cells. Further tests showed that selenium nanoparticles treatment leads to apoptotic cell death enhancement and glucose metabolism reduction, and this process was in a reactive oxygen species pathway-dependent manner. These results may provide a novel direction for glioma therapy in the future.

Effects of superficial temporal artery-middle cerebral artery bypass on hemodynamics and clinical outcomes in the patients with atherosclerotic stenosis in the intracranial segment of internal carotid artery and middle cerebral artery.

OBJECTIVE: To observe the effects of superficial temporal artery-middle cerebral artery bypass (STA-MCA bypass) on hemodynamics and clinical outcomes in the patients with atherosclerotic stenosis in the intracranial segment of internal carotid artery and (or) middle cerebral artery. PATIENTS AND METHODS: The data of 63 patients who had the symptoms of cerebral ischemia in recent 3 months, intracranial segment of internal carotid artery (ISICA) and (or) middle cerebral artery (MCA) stenoses or occlusion showed by digital subtraction angiography (DSA), and reduced cerebral perfusion displayed by CT perfusion (CTP) imaging were retrospectively collected in this study. According to the patient's choice of different treatment methods (STA-MCA bypass and drugs), these patients were allocated into two groups: Bypass group (30 cases) and Drug group (33 cases). Postoperative symptoms, anastomotic patency and hemodynamics were observed in the Bypass group. Post-treatment ischemic events and clinical outcomes were recorded in the two groups and were compared between the two groups. RESULTS: In the Bypass group, DSA all showed anastomotic patency in 28 patients (93.3%, 28/30), and the improvement rate of CTP was all significantly higher in the patients with stage-III CTP than in the patients with stage-II CTP at post-operative 3 days and 6 months (95% vs 60%). Post-treatment ischemic event incidence (13.3% vs 48.5%) and annual stroke rate (6.7% vs 25.6%) were significantly lower in the Bypass group than in the Drug group (All P < 0.05). Pre-treatment National Institutes of Health Stroke Scale (NIHSS) score and Modified Rankin Scale (MRS) score were not significantly different between the two groups, but the NIHSS (2.87±0.19 and 2.4±0.19 vs 4.03±0.47 and 3.97±0.49) and MRS (1.13±0.09 and 1.0±0.07 vs 1.55±0.14 and 1.52±0.15) all were significantly lower in the Bypass group than in the Drug group at post-treatment 6 and 24 months (all P < 0.05). CONCLUSION: STA-MCA bypass can improve cerebral blood perfusion and reduce the incidence of stroke in the patients who have ISICA and (or) MCA-related symptoms, 70%-100% of stenosis, and above stage-ⅠCTP. However, this conclusion remains to be further confirmed.

Carotid-Carotid Artery Crossover Bypass with a Synthetic Vascular Graft for Symptomatic Type 1A Common Carotid Artery Occlusion.

OBJECTIVE: To investigate the effect of carotid-carotid artery crossover bypass with a synthetic vascular graft for symptomatic type 1A common carotid artery occlusion (CCAO). METHODS: A retrospective analysis was conducted of patients with symptomatic type 1A CCAO who underwent carotid-carotid artery crossover bypass surgery via a retropharyngeal route with a synthetic vascular graft in the Department of Neurosurgery at our hospital. Preoperative demographic data, surgical complications, incidence of stroke during follow-up, and other clinical data were summarized. RESULTS: Between 2011 and 2016, carotid-carotid artery crossover bypass was performed with a synthetic vascular graft in 4 patients with type 1A CCAO. The mean patient age was 63.3 years (range, 49-69 years). Clinical symptoms included dizziness, amaurosis fugax, persistent limb numbness, and transient ischemic attack. In all 4 patients, postoperative computed tomography angiography showed internal carotid artery thickening due to successful bypass, whereas computed tomography perfusion showed improved postoperative cerebral perfusion on the side of the lesion. The sole perioperative complication was a complaint of foreign body sensation on swallowing in 1 patient. The mean duration of follow-up was 40.3 months (range, 14-77 months), during which no newly occurred cerebral ischemia or synthetic vascular graft occlusion was observed. CONCLUSIONS: Carotid-carotid artery crossover bypass with a synthetic vascular graft is a safe and effective therapeutic approach for patients with symptomatic type 1A CCAO. However, studies with larger series are needed to enable more precise conclusions.

NF-κB induces miR-148a to sustain TGF-ß/Smad signaling activation in glioblastoma.

BACKGROUND: Inflammatory cytokines and transforming growth factor-ß (TGF-ß) are mutually inhibitory. However, hyperactivation of nuclear factor-κB (NF-κB) and TGF-ß signaling both emerge in glioblastoma. Here, we report microRNA-148a (miR-148a) overexpression in glioblastoma and that miR-148a directly suppressed Quaking (QKI), a negative regulator of TGF-ß signaling. METHODS: We determined NF-κB and TGF-ß/Smad signaling activity using pNF-κB-luc, pSMAD-luc, and control plasmids. The association between an RNA-induced silencing complex and QKI, mitogen-inducible gene 6 (MIG6), S-phase kinase-associated protein 1 (SKP1), and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) mRNA was tested with microribonucleoprotein immunoprecipitation and real-time PCR. Xenograft tumors were established in the brains of nude mice. RESULTS: QKI suppression induced an aggressive phenotype of glioblastoma cells both in vitro and in vivo. Interestingly, we found that NF-κB induced miR-148a expression, leading to enhanced-strength and prolonged-duration TGF-ß/Smad signaling. Notably, these findings were consistent with the significant correlation between miR-148a levels with NF-κB hyperactivation and activated TGF-ß/Smad signaling in a cohort of human glioblastoma specimens. CONCLUSIONS: These findings uncover a plausible mechanism for NF-κB-sustained TGF-ß/Smad activation via miR-148a in glioblastoma, and may suggest a new target for clinical intervention in human cancer.

Clipping of anterior communicating artery aneurysms in the early post-rupture stage via transorbital keyhole approach--Chinese neurosurgical experience.

The anterior communicating artery (AComA) complex is the site at which intracranial aneurysms occur most frequently. At present, effective treatments for AComA aneurysms are yet to be developed. Here, we present our experience in successfully managing AComA aneurysms via the transorbital keyhole approach. A total of 52 patients having a history of aneurysm rupture received surgery. All patients were assigned a Hunt-Hess grade prior to surgery. The cistern was opened to expose the AComA complex using a keyhole approach, and aneurysms were then surgically clipped with the assistance of neuroendoscopy or indocyanine green angiography. Surgery outcomes were confirmed using computed tomography angiography (CTA). Each of the 52 AComA aneurysms was successfully clipped with a single operation. Three of these patients experienced intraoperative aneurysm rupture. Five had postoperative hydrocephalus which was successfully treated with ventriculoperitoneal shunt. All patients survived the surgical procedure. Using the Glasgow Outcome Scale scores for evaluation, 39 patients (75.0%) had good recovery, 9 (17.3%) had moderate disability, 2 (3.8%) had severe disability, and 2 patients who had been in preoperative comas (3.8%) remained in a vegetative state. During the follow-up period, CTA showed no recurrence of rupture or bleeding in all cases. Results of logistic analysis indicated that the transorbital keyhole approach was feasible based on the patients' preoperative Hunt-Hess grades, which should be considered a priority in using this approach in the treatment of ruptured AComA aneurysms.

Construction and identification of human glial cell-derived neurotrophic factor gene-modified Schwann cells from rhesus monkeys.

The objective of this study was to construct stable rhesus monkey Schwann cells (SCs) modified with the human glial cell-derived neurotrophic factor (hGDNF) gene. hGDNF gene amplification was performed with pUC19-hGDNF as templates, and then the coding sequence of hGDNF was inserted into the eukaryotic expression vector pBABE-puro to obtain the recombinant vector pBABE-puro-hGDNF. The recombinant vector pBABE-puro-hGDNF was identified with restriction enzyme, and then underwent DNA sequencing. SCs from rhesus monkeys were transfected with the recombinant vector pBABE-puro-hGDNF, and then the expression levels of mRNA and protein of the hGDNF gene were determined with real-time fluorescence quantitative PCR and Western blot, respectively, in the transfected SCs. The biological activity of GDNF gene-modified SCs (GDNF-SCs) was assessed by MTT assay. The length of the hGDNF coding sequence of PCR products was 569 bp. After the recombinant eukaryotic expression vectors were digested with restriction enzyme, there was a specific segment of 596 bp. The results of DNA sequencing of the specific segment of 596 bp were the same as that of hGDNF in GenBank, suggesting that the hGDNF gene was successfully inserted into the recombinant retrovirus vectors. The expression levels of mRNA and protein were significantly higher in transfected SCs as compared to nontransfected SCs (p<0.05). MTT assay indicated that the OD value was significantly higher in GDNF-SCs group than in SCs and DMEM groups (p<0.05). hGDNF-SCs can steadily and efficiently release hGDNF. This study provides a basis for cell therapy of nerve injury.

[Construction and identification of rhesus monkey Schwann cells modified with human glial cell derived neurotrophic factor gene].

OBJECTIVE: To construct the rhesus monkey Schwann cells (SCs) modified with human glial cell derived neurotrophic factor (hGDNF) gene. METHODS: The coding sequence ofhGDNF amplified by PCR from pUC19-hGDNF was inserted into eukaryotic expression vector pBABE-puro. The recombinant eukaryotic expression vector pBABE-puro-hGDNF was identified with restriction enzyme digestion and DNA sequencing. The SCs were isolated from rhesus monkeys, cultured and purified. The SCs were transfected with the recombinant retrovirus vector containing hGDNF gene. The mRNA and protein expressions of hGDNF were analyzed by real-time fluorescent quantitative PCR and Western blot. RESULTS: The PCR product of hGDNF coding sequence was a 596 bp specific segment. The recombinant eukaryotic expression vector was digested into a 596 bp specific segment by specific restriction enzyme and another segment. The 596 bp segment confirmed by DNA sequencing was consistent with hGDNF sequence on GenBank. Restriction enzyme digestion and sequencing results showed that the coding sequence of hGDNF was successfully inserted into the recombinant retrovirus vector and the mRNA and protein expressions of hGDNF were significantly higher in transfected SCs than non-transfected SCs (P < 0.05). CONCLUSION: The rhesus monkey SCs modified with hGDNF gene are successfully constructed and hGDNF can be released continuously and stably, which will provide a foundation for the further research about cell therapy of hGDNF-SCs in the repair of injured nerve.