Integrated Bioinformatic Analysis and Validation Identifies Immune Microenvironment-Related Potential Biomarkers in Alzheimer's Disease.

BACKGROUND: Alzheimer's disease (AD) is one of the most common neurodegenerative diseases, accompanied by cognitive and memory impairment, accounting for about 60% - 80% of dementia types. The pathogenesis of AD has not been clarified, and there is no effective therapy to prevent or treat AD. In this study, we aimed to identify the potential biomarkers involved in the brain immune microenvironment in AD. METHODS: AD datasets from GEO database were obtained to identify the differentially expressed disease-related genes (DEDRGs) in AD through weighted gene co-expression network analysis (WGCNA) and differential expression analysis. Functional Enrichment analysis was performed to explore the potential biological function of DEDRGs. The hub DEDRGs were identified through the protein-protein interaction (PPI) network. Furthermore, the CIBERSORT algorithm was employed to bulk gene expression profiles of AD to depict the immune microenvironment characteristics in AD. Pearson's correlation analysis was utilized to depict the correlation between each of immune cells and hub DEDRGs. RESULTS: A total of 27 DEDRGs were identified through WGCNA and differential expression analysis. Functional enrichment analysis of 27 DEDRGs indicated that chemokine signaling pathway was the most significantly enriched KEGG pathway, response to biotic stimulus was the most significantly enriched GO term, and most of DEDRGs were enriched into urinary system cancer in DO analysis. 6 hub DEDRGs, ANGPT1, CCL2, CD44, CXCR4, GJA1 and VCAM1, were screened through PPI network and all of them were up-regulated in AD. Immune infiltration analysis revealed that there were higher infiltration levels of T cells CD4 memory activated, T cells gamma delta, NK cells resting and macrophages M0, and lower infiltration level of NK cell activated in AD, and macrophages M2 owned the highest positively association with VCAM1 and CXCR4, but VCAM1 was statistically and negatively correlated to T cells CD8. CONCLUSION: Our study identified 6 hub DEDRGs, ANGPT1, CCL2, CD44, CXCR4, GJA1 and VCAM1, were statistically associated with immune infiltrating cells, and were significantly related to the pathological development of AD, which may provide a theoretical basis for developing potential biomarkers and implementing effective therapies against AD.

Polydopamine deposition with anodic oxidation for better connective tissue attachment to transmucosal implants.

BACKGROUND AND OBJECTIVE: Nowadays, most designs for the transmucosal surface of implants are machined-smooth. However, connective tissue adhered to the smooth surface of an implant has poor mechanical resistance, which can render separation of tissue from the implant interface and induce epithelial downgrowth. Modification of the transmucosal surface of implants, which can help form a good seal of connective tissue, is therefore desired. We hypothesized that anodic oxidation (AO) and polydopamine (PD) deposition could be used to enhance the attachment between an implant and peri-implant connective tissue. We tested this hypothesis in the mandibles of Beagle dogs. MATERIAL AND METHODS: AO and PD were used to modify the transmucosal region of transmucosal implants (implant neck). The surface microstructure, surface roughness and elemental composition were investigated in vitro. L929 mouse fibroblasts were cultured to test the effect of PD on cell adhesion. Six Beagle dogs were used for the in vivo experiment (n = 6 dogs per group). Three months after building the edentulous animal model, four groups of implants (control, AO, PD and AO + PD) were inserted. After 4 months of healing, samples were harvested for histometric analyses. RESULTS: The surfaces of anodized implant necks were overlaid with densely distributed pores, 2-7 µm in size. On the PD-modified surfaces, N1s, the chemical bond of nitrogen in PD, was detected using X-ray photoelectron spectroscopy. L929 developed pseudopods more quickly on the PD-modified surfaces than on the surfaces of the control group. The in vivo experiment showed a longer connective tissue seal and a more coronally located peri-implant soft-tissue attachment in the AO + PD group than in the control group (P < .05). CONCLUSION: The modification of AO + PD on the implant neck yielded better attachment between the implant and peri-implant connective tissue.

Identification of the novel allele, HLA-DRB1*09:30, by sequence-based high resolution typing.

HLA-DRB1*09:30 shows a single nucleotide difference from DRB1*09:01:02 at nucleotide 250 (CGG→TGG).

α5ß1-Integrin promotes tension-dependent mammary epithelial cell invasion by engaging the fibronectin synergy site.

Tumors are fibrotic and characterized by abundant, remodeled, and cross-linked collagen that stiffens the extracellular matrix stroma. The stiffened collagenous stroma fosters malignant transformation of the tissue by increasing tumor cell tension to promote focal adhesion formation and potentiate growth factor receptor signaling through kinase. Importantly, collagen cross-linking requires fibronectin (FN). Fibrotic tumors contain abundant FN, and tumor cells frequently up-regulate the FN receptor α5ß1 integrin. Using transgenic and xenograft models and tunable two- and three-dimensional substrates, we show that FN-bound α5ß1 integrin promotes tension-dependent malignant transformation through engagement of the synergy site that enhances integrin adhesion force. We determined that ligation of the synergy site of FN permits tumor cells to engage a zyxin-stabilized, vinculin-linked scaffold that facilitates nucleation of phosphatidylinositol (3,4,5)-triphosphate at the plasma membrane to enhance phosphoinositide 3-kinase (PI3K)-dependent tumor cell invasion. The data explain why rigid collagen fibrils potentiate PI3K activation to promote malignancy and offer a perspective regarding the consistent up-regulation of α5ß1 integrin and FN in many tumors and their correlation with cancer aggression.

Detection of a novel HLA-B allele, HLA-B*39:119, in a Chinese individual.

HLA-B*39:119 allele differs from HLA-B*39:01:01:01 by a single nucleotide substitution at position 488.

A study of HLA-B*15:02 in 9 different Chinese ethnics: Implications for carbamazepine related SJS/TEN.

BACKGROUND: HLA-B*15:02 is a known biomarker for carbamazepine (CBZ)-induced Stevens-Johnson syndrome and toxic epidermal necrolysis (SJS/TEN) in some ethnic populations. The US FDA recommends B*15:02 screening for Asian and other populations with a high prevalence of B*15:02 prior to treatment with CBZ to prevent drug-related SJS/TEN. MATERIALS AND METHODS: A total of 1607 blood samples were collected from volunteer blood donors who were ethnic minorities living in the Yunnan province of southwestern China, including 153 Yi, 193 Naxi, 167 Miao, 156 Lisu, 166 Derung, 211 Bai, 184 Hani, 198 Dai, and 179 Zhuang. The genetic diversity of the HLA-B*15:02 genes in the ethnic minority samples was examined using sequence based typing at high resolution. RESULTS: The allele frequencies of HLA-B*15:02 in the Yi, Naxi, Miao, Lisu, Derung, Bai, Hani, Dai, and Zhuang populations were 4.25%, 4.4%, 5.09%, 5.77%, 6.33%, 7.82%, 8.15%, 9.6%, and 15.36%, respectively. The frequencies of HLA-B*15:02 carriers in the Yi, Naxi, Miao, Lisu, Derung, Bai, Hani, Dai, and Zhuang populations were 8.5%, 8.8%, 9.58%, 10.9%, 12.65%, 15.64%, 16.3%, 18.69%, and 28.49%, respectively. CONCLUSION: The HLA-B*15:02 allele frequencies indicated that the prevalence of B*15:02 was different among the different ethnic populations. Because the number of carriers of B*15:02 was high in some ethnic populations, larger studies are required to confirm these findings. The Zhuang population had the highest frequency of B*15:02 in this study. More attention should be paid to CBZ-induced SJS/TEN in Chinese minority populations.

Analysis of postcytokinetic roles of cytokinetic components in Caenorhabditis elegans.

Cytokinesis requires the interplay between the cytoskeleton and plasma membrane. Emerging evidence indicates that some cytokinetic components are essential for the postcytokinetic events such as epithelium organization and neural development. We have recently developed live cell imaging and conditional knockout techniques to visualize cytokinetic proteins in Caenorhabditis elegans Q neuroblasts and separate their postcytokinetic functions from cytokinetic ones. Here we describe how the fluorescent reporter strains and conditional knockout C. elegans are generated and how live cell imaging of Q neuroblast development are performed in our laboratory. Using these protocols, we uncovered a novel role of Anillin in stabilizing the actin network during neuronal migration and neurite outgrowth, and the postcytokinetic fate of midbody, which is released into the extracellular space and degraded by the adjacent macrophage using an apoptotic mimicry. These protocols could also be applicable to study other cellular processes in C. elegans or adapted to other model organisms, to provide better insights into their developmental basis.

HLA-B*46:40:02, a novel HLA-B*46 allele identified in a Chinese individual by sequence-based typing.

HLA-B*46:40:02 has one synonymous nucleotide change from HLA-B*46:40:01 in codon 23, exon 2 (ATT>ATC).

Identification of a novel HLA-A allele, A*11:209 by sequence-based high-resolution typing in an HIV-infected individual.

The novel allele A*11:209 shows three nucleotides difference from A*11:27 in exon 3.

Radiotherapy effects on brain/bone metastatic adenocarcinoma lung cancer and the importance of EGFR mutation test.

This study proposed to retrospectively analyze the efficacy of radiotherapy on brain/bone metastases in patients with stage IV lung adenocarcinoma and to evaluate the correlation between overall survival after radiotherapy and other factors including metastatic sites and EGFR mutation status. 115 patients with Stage IV lung adenocarcinoma admitted to our center from March, 2011 to December, 2013 were enrolled. They presented with metastases to no other solid organs except the bone or brain and had received no prior treatment. 50 patients received EGFR mutation test with 32 detected as EGFR mutant and 18 wild-type. Patients with brain metastases were treated with 40 Gy whole brain irradiation (WBI) in 2 Gy fractions; patients with bone metastases were treated with 30 Gy local irradiation in 3 Gy fractions or 40 Gy in 2 Gy fractions. All the patients received systemic therapy during or after radiotherapy and 68 received targeted therapy.The median overall survival of patients with solitary brain metastases, solitary bone metastases or combined brain and bone metastases were 8.50 months, 8.50 months and 9.50 months respectively, revealing no significant difference (p=0.57). The median overall survival of patients with EGFR mutations was 10.25 months, longer than the 8.75 months of patients without EGFR mutations, revealing no significant difference (p=0.57). The median overall survival of EGFR mutant patients with solitary bone metastases, solitary brain metastases or combined brain and bone metastases were 7.50 months, 10.50 months and 11.50 months respectively, revealing no significant difference (p=0.91). 36 patients with untested EGFR mutation status received EGFR-TKI. Among EGFR mutant patients, 10 didn't receive targeted therapy; 8 were administered Erlotinib and 14 Gefitinib with median overall survival of 10.25 months and 14.5 months, showing no significant difference (p=0.11) between the two drugs. When patients with stage IV lung adenocarcinoma have been treated by early radiotherapy, the overall survival doesn't correlate with metastatic sites. Radiotherapy could extend survival for EGFR mutant patients with stage IV lung adenocarcinoma. EGFR mutation test should be performed before treatment of the disease.

In vitro subminimum inhibitory concentrations of macrolide antibiotics induce macrolide resistance in Mycoplasma pneumoniae.

AIM: This study aims to investigate the inducing effect of subminimum inhibitory concentrations of macrolide antibiotics on Mycoplasma pneumoniae (M. pneumoniae) resistance to drugs. MATERIALS AND METHODS: One M. pneumoniae reference strain M129 (ATCC 29342) and 104 clinical isolates were incubated at 37C for 6-8 days. Genomic DNA of M. pneumoniae was extracted using TIANamp Bacteria DNA kit and amplified by polymerase chain reaction (PCR). RESULTS: Ten sensitive isolates obtained from 104 M. pneumoniae clinical isolates were induced by subminimum inhibitory concentrations of macrolide antibiotics. Among them, three were found to possess mutations in L4 and L22 ribosomal proteins. Two cases carried simultaneously the C162A and A430G mutations of L4 and the T279C mutation of L22. In addition, one case had only the A209T mutation of L4. CONCLUSIONS: Repeated in vitro exposure to subminimum inhibitory concentrations of macrolide antibiotics could induce selective mutations in ribosomal genes of M. pneumoniae clinical isolates that cause resistance to macrolide antibiotics. Hippokratia 2015, 19 (1): 57-62.

The new allele HLA-DPB1*363:01 identified by sequence-based typing in a donor from the Chinese Marrow Donor Program.

The new allele HLA-DPB1*363:01 most closely resembles DPB1*92:01, differing at a single position 191 (exon 2, codon 35).

HLA-B*58:63, a novel allele identified by sequence-based typing in a Chinese bone marrow voluntary donor.

HLA-B*58:63 differs from HLA-B*58:01:01 by one nucleotide at position 248 resulting in a single amino acid change.

Identification of a novel HLA-A*24 allele, A*24:289, in a Chinese individual.

The new allele, A*24:289 is different to A*24:03:01 at codons 151, 152 and 156 at exon 3.

A novel HLA-B allele, HLA-B*15:325, is identified by sequence-based typing in a HIV infected individual.

The novel allele B*15:325 shows difference from B*15:02:01 at codon 127 resulting in changes from Asn to Ser.

Comparison of 16S rRNA gene PCR and blood culture for diagnosis of neonatal sepsis.

UNLABELLED: Septicemia is a common cause of morbidity and mortality among newborns in the developing world. However, accurate clinical diagnosis of neonatal sepsis is often difficult because symptoms and signs are often nonspecific. Blood culture has been the gold standard for confirmation of the diagnosis. However, the sensitivity is low and results are usually not promptly obtained. Therefore, the diagnosis of sepsis is often based on clinical signs in association with laboratory tests such as platelets count, immature/total neutrophils ratio (I/T), and a rise in C-reactive protein (CRP). Polymerase chain reaction (PCR) methods for the detection of neonatal sepsis represent new diagnostic tools for the early identification of pathogens. METHODS: During a 4-month prospective study, 16S rRNA PCR was compared with conventional blood culture for the diagnosis of neonatal bacterial sepsis. In addition, the relationship between known risk factors, clinical signs, laboratory parameters, and the diagnosis of sepsis was considered. RESULTS: Sepsis was suspected in 706 infants from the intensive neonatal care unit. They all were included in the study. The number of positive cultures and positive PCR results were 95 (13.5%) and 123 (17.4%), respectively. Compared with blood culture, the diagnosis of bacterial sepsis by PCR revealed a 100.0% sensitivity, 95.4% specificity, 77.2% positive predictive value, and 100.0% negative predictive value. In this study, Apgar scores at 5 min, weight, icterus, irritability, feeding difficulties, gestational age (GA), premature rupture of membrane (PRM), platelets count, I/T, and a marked rise in CRP were important in establishing the diagnosis of sepsis in the newborn. In addition, weight, GA, PRM, irritability, duration of antibiotic usage, mortality rate, and number of purulent meningitis cases were significantly different between early-onset sepsis and late-onset sepsis. CONCLUSION: 16S rRNA PCR increased the sensitivity in detecting bacterial DNA in newborns with signs of sepsis, allowed a rapid detection of the pathogens, and led to shorter antibiotic courses. However, uncertainty about the bacterial cause of sepsis was not reduced by this method. 16S rRNA PCR needs to be further developed and improved. Blood culture is currently irreplaceable, since pure isolates are essential for antimicrobial drug susceptibility testing.

Involvement of decreased hypoxia-inducible factor 1 activity and resultant G1-S cell cycle transition in radioresistance of perinecrotic tumor cells.

Cancer patients often suffer from local tumor recurrence after radiation therapy. Some intracellular and extracellular factors, such as activity of hypoxia-inducible factor 1 (HIF-1), cell cycle status and oxygen availability, have been suggested to affect DNA damage responses and eventual radioresistant characteristics of cancer cells. But when, where, and how these factors affect one another and induce cellular radioresistance is largely unknown. Here, we analyzed mechanistic and spatio-temporal relationships among them in highly heterogeneous tumor microenvironments. Experiments in vitro demonstrated that a decrease in the glucose concentration reduced the transcriptional activity of HIF-1 and expression of a downstream gene for the cell cycle regulator p27(Kip1) even under hypoxic conditions. Then, the proportion of cells in the radioresistant S phase increased, whereas that in the radiosensitive G1 phase decreased, significantly. Immunohistochemical analyses showed that cancer cells in perinecrotic hypoxic regions, which should be under low-glucose conditions, expressed little HIF-1α, and therefore, were mainly in S phase and less damaged by radiation treatment. Continuous administration of glucagon, which increases the blood glucose concentration and so improves glucose availability in perinecrotic hypoxic regions, induced HIF-1α expression and increased radiation-induced DNA damage. Taken all together, these results indicate that cancer cells in perinecrotic regions, which would be under low-glucose and hypoxic conditions, obtain radioresistance by decreasing the level of both HIF-1 activity and p27(Kip1) expression, and adjusting their cell cycle to the radioresistant S phase.

Prevalence of post-traumatic stress disorder among adolescents after the Wenchuan earthquake in China.

BACKGROUND: The Wenchuan earthquake was a catastrophic earthquake in China. The aim of this study is to explore longitudinally the rates of post-traumatic stress disorder (PTSD) and depression in adolescents after the Wenchuan earthquake, and to identify independent predictors of PTSD. METHOD: PTSD and depression symptoms among adolescents at 6, 12 and 18 months after the Wenchuan earthquake were investigated using the PTSD Checklist Civilian Version and the Beck Depression Inventory (BDI). Subjects in this study included 548 high school student survivors in a local boarding high school. RESULTS: The rates of PTSD symptoms were 9.7%, 1.3% and 1.6% at the 6-, 12- and 18-month follow-ups, respectively. BDI scores were found to be the best predictor of severity of PTSD at 6, 12 and 18 months. Gender was another variable contributing significantly to PTSD at 6 and 12 months after the earthquake. In the 12-month follow-up, home damage was found to be a predictor of severity of PTSD symptoms. Being a child with siblings was found to be a predictor of severity of PTSD symptoms at 12 and 18 months after the earthquake. CONCLUSIONS: PTSD symptoms changed gradually at various stages after the earthquake. Depression symptoms were predictive of PTSD symptoms in the 18-month follow-up study. Other predictors of PTSD symptoms included female gender and being a child with siblings. The results of this study may be helpful for further mental health interventions for adolescents after earthquakes.

Apolipoprotein A5 polymorphisms and risk of coronary artery disease: a meta-analysis.

The relation has not been reported consistently between the polymorphisms in the gene of apolipoprotein A5 (APO A5) and coronary artery disease (CAD). To clarify the discrepancy, we conducted a comprehensive search of PubMed and EMBASE for all available casecontrol studies to explore the association between two APO A5 polymorphisms and CAD. Two reviewers independently selected studies. Statistical analyses were carried out using the STATA software package v 10.0. Thirteen studies investigated the association between the APO A5 -1131T>C polymorphism and risk of CAD were selected in this meta-analysis with 5,050 cases and 7,272 controls. For the S19W APO A5 gene polymorphism, 5 studies were included with 2,196 cases and 3,933 controls. We observed a significant statistical association between Apo A5 -1131T>C polymorphism and CAD (recessive genetic model: OR = 1.73, 95% CI = 1.37-2.19; dominant genetic model: OR = 1.42, 95% CI = 1.25-1.61; allelic contrast: OR = 1.31, 95% CI = 1.22-1.39, respectively). After restricting our analysis to Chinese individuals, we found that the association was stronger. We also observed strong association between the APO A5 S19>W polymorphism and risk of CAD under a recessive genetic model. This meta-analysis reveals that the minor allele of the -1131T>C polymorphism in the promoter of APO A5 gene significantly increases the susceptibility to CAD. This effect is more pronounced in Chinese subjects.

Clinical observation of 73 nasopharyngeal carcinoma patients treated by helical tomotherapy: the China experience.

The preliminary short-term clinical outcome of 73 nasopharyngeal carcinoma (NPC) patients treated with helical tomotherapy at our cancer institute has been evaluated. Between September 2007 and September 2009, 73 newly diagnosed NPC patients were treated with helical tomotherapy. The distributions of clinical stages according to the UICC 2002 Staging System were: 6, 27, 24, and 16 for Stage I, IIa-b, III, and IVa-b, respectively. The prescription dose was 70-74 Gy/33F to planning gross tumor volume containing the primary tumor and positive lymph nodes, with 60-62.7 Gy/33F to high risk planning target volume, while delivering 52-56 Gy/33F to low risk planning target volume. Twenty-four patients were treated with radiation therapy as single modality, 25 with concurrent cisplatin-based chemotherapy with or without anti-EGFR monoclonal antibody therapy, and 24 with concurrent anti-EGFR monoclonal antibody therapy. Setup errors were analyzed. Side-effects were evaluated with the established RTOG/EORTC criteria. Average beam-on-time was 468.8 sec/F (396.7-696.1 sec). The setup errors in the lateral, longitudinal and vertical directions were 0.00 ± 1.79 mm, -0.55± 2.17 mm and 0.38 ± 1.43 mm, corresponding to 3.80 mm, 4.20 mm, and 2.46 mm as the CTV-PTV margin in these directions. The grade 0, 1, 2 and 3 acute skin toxicity was 2.7%, 76.7%, 13.8% and 6.8%; the grade 0, 1, 2 and 3 acute mucositis was 1.4%, 32.9%, 60.2% and 5.5%; and the grade 0, 1, 2 and 3 acute xerostomia was 4.0%, 45.3%, 50.7% and 0, respectively. Only 5 patients suffered from grade 3 or 4 leucopenia. Xerostomia resolved with passing of time and no grade 2 or more xerostomia was noted one year after radiation therapy. Concurrent chemotherapy significantly increased incidence of severe acute toxicities. One month after radiation therapy the remission rates of primary tumor and positive lymph nodes were 91.8% and 98.1%, respectively. The median follow-up was 14.8 months. The one-year relapse-free survival, distant metastasis-free survival and overall survival was 95.6%, 97.2% and 94.8%, respectively. In conclusion, the incidence of severe acute toxicities and late xerostomia was relatively infrequent for NPC patients treated with helical tomotherapy. The long-term clinical outcome for these patients is under investigation.