Airway eosinophilic inflammation is a pivotal feature of asthma. Epithelial cells play critical roles in airway eosinophilia. We hypothesized that epithelial microRNAs (miRNAs) are involved in airway eosinophilia.

This study investigated the associations between epithelial and plasma miR-181b-5p and airway eosinophilic inflammation, and the possible mechanism by which miR-181b-5p participates in eosinophilic inflammation.

Epithelial miRNAs expression was profiled by miRNA array in eight subjects with asthma and four healthy controls. Epithelial miR-181b-5p expression was confirmed by quantitative PCR in the subjects for array experiment and another cohort including 21 subjects with asthma and 10 controls. Plasma miR-181b-5p was determined by quantitative PCR in 72 subjects with asthma and 35 controls. Correlation assays between epithelial and plasma miR-181b-5p expression and airway eosinophilia were performed. The target of miR-181b-5p, SPP1, was predicted by online algorithms and verified in BEAS-2B cells. The role of miR-181b-5p in epithelial proinflammatory cytokine expression was examined in an in vitro system.

Epithelial miR-181b-5p expression was decreased in subjects with asthma. Epithelial miR-181b-5p levels were inversely correlated with sputum and bronchial submucosal eosinophilia. Plasma miR-181b-5p was decreased and correlated with epithelial miR-181b-5p in subjects with asthma. There was a strong inverse correlation between plasma miR-181b-5p and airway eosinophilia in subjects with asthma. Plasma miR-181b-5p was increased after inhaled corticosteroids treatment. We verified that SPP1 is a target of miR-181b-5p. In human bronchial epithelial cells, miR-181b-5p regulated IL-13-induced IL-1β and CCL11 expression by targeting SPP1. Dexamethasone restored IL-13-induced miR-181b-5p down-regulation and suppressed IL-13-induced SPP1, IL-1β and CCL11 expression.

Epithelial and plasma miR-181b-5p are potential biomarkers for airway eosinophilia in asthma. MiR-181b-5p may participate in eosinophilic airway inflammation by regulating proinflammatory cytokines expression via targeting SPP1.

Second-generation irreversible epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI), afatinib, has been approved for treating EGFR mutant lung cancer patients, but the mechanism of acquired resistance to afatinib has not been well studied. In this study, we established afatinib acquired resistant cell lines. Gene array technology was used to screen changes in gene expression between afatinib-resistant lung cancer cells and parental cells. Our results showed that secreted phosphoprotein 1 (SPP1) was significantly increased in afatinib-resistant lung cancer cells. To study the effect of SPP1 on afatinib resistance, siSPP1 was used to knock down SSP1 in afatinib-resistant lung cancer cells. Then sensitivity to afatinib and invasive ability were studied. We found that knockdown of SPP1 increased sensitivity of lung cancer cells to afatinib and decrease the ability of invasion. Of clinical significance, we found that SSP1 was upregulated in lung cancer tissues compared with adjacent normal tissues, and low level of SSP1 was strongly associated with better overall survival. Our results suggest that SPP1 enhanced the second-generation EGFR TKI resistance in lung cancer, and inhibiting SPP1 might be a therapeutic target to overcome afatinib resistance.

Follicle development is accompanied by proliferation of granulosa cells and increasing oocyte size. To obtain high-quality oocytes in vitro, it is important to understand the processes that occur in oocytes and granulosa cells during follicle development and the differences between in vivo and in vitro follicle development. In the present study, oocytes and granulosa cells were collected from early antral follicles (EAFs, 0.5-0.7 mm in diameter), small antral follicles (SAFs, 1-3 mm in diameter), large antral follicles (LAFs, 3-7 mm in diameter), and in vitro grown oocyte-and-granulosa cell complexes (OGCs), which were cultured for 14 days after collection from EAFs. Gene expression was analyzed comprehensively using the next-generation sequencing technology. We found top upstream regulators during the in vivo follicle development and compared them with those in in vitro developed OGCs. The comparison revealed that HIF1 is among the top regulators during both in vivo and in vitro development of OGCs. In addition, we found that HIF1-mediated upregulation of glycolysis in granulosa cells is important for the growth of OGCs, but the cellular metabolism differs between in vitro and in vivo grown OGCs. Furthermore, on the basis of comparison of upstream regulators between in vivo and in vitro development of OGCs, we believe that low expression levels of FLT1 (VEGFA receptor), SPP1, and PCSK6 can be considered causal factors of the suboptimal development under in vitro culture conditions.

OBJECTIVE

Cell adhesion plays important roles in maintaining the structural integrity of connective tissues and sensing changes in the biomechanical environment of cells. The objective of the present investigation was to extend our understanding of the effect of cyclic mechanical strain on the expression of adhesion-related genes by human periodontal ligament cells.

METHODS

Cultured periodontal ligament cells were subjected to a cyclic in-plane tensile deformation of 12% for 5 s (0.2 Hz) every 90 s for 6-24 h in a Flexercell FX-4000 Strain Unit. The following parameters were measured: (i) cell viability by the MTT assay; (ii) caspase-3 and -7 activity; and (iii) the expression of 84 genes encoding adhesion-related molecules using real-time RT-PCR microarrays.

RESULTS

Mechanical stress reduced the metabolic activity of deformed cells at 6 h, and caspase-3 and -7 activity at 6 and 12 h. Seventy-three genes were detected at critical threshold values < 35. Fifteen showed a significant change in relative expression: five cell adhesion molecules (ICAM1, ITGA3, ITGA6, ITGA8 and NCAM1), three collagen α-chains (COL6A1, COL8A1 and COL11A1), four MMPs (ADAMTS1, MMP8, MMP11 and MMP15), plus CTGF, SPP1 and VTN. Four genes were upregulated (ADAMTS1, CTGF, ICAM1 and SPP1) and 11 downregulated, with the range extending from a 1.76-fold induction of SPP1 at 12 h to a 2.49-fold downregulation of COL11A1 at 24 h.

CONCLUSIONS

The study has identified several mechanoresponsive adhesion-related genes, and shown that onset of mechanical stress was followed by a transient reduction in overall cellular activity, including the expression of two apoptosis 'executioner' caspases.

BACKGROUND

Secreted phosphoprotein 1 (SPP1 or Osteopontin, OPN) is a multifunctional matricellular glycoprotein involved in development and regeneration of skeletal muscle. Previously, we have demonstrated that porcine SPP1 shows breed-related differential mRNA expression during myogenesis. With the aim to identify putative contributing cis-regulatory DNA variation we resequenced the 5' upstream region of the gene in the respective breeds Pietrain and Duroc. We found two single nucleotide polymorphisms (SNP; [GenBank:M84121]: g.1804C>T and g.3836A>G). We focused our investigation on the SNP g.3836A>G, because in silico analysis and knowledge about the regulation of SPP1 suggested an effect of this SNP on a CCAAT/enhancer binding protein beta (C/EBPbeta) responsive transcriptional enhancer.

RESULTS

Using electrophoretic mobility shift assay we demonstrated that, similar to human SPP1, the 3' terminal end of the first intron of porcine SPP1 harbors a C/EBPbeta binding site and showed that this binding site is negatively affected by the mutant G allele. Genotyping of 48 fetuses per breed revealed that the G allele segregated exclusively in Duroc fetuses with a frequency of 57 percent. Using real-time quantitative PCR we showed that, consistent with its negative effect on a transcriptional enhancer element, the G allele tends to decrease mRNA abundance of SPP1 in the fetal musculus longissimus dorsi (approximately 1.3 fold; P>> or = 0.1). Moreover, we showed that the SNP g.3836A>G leads to ubiquitous aberrant splicing of the first intron by generating a de novo and activating a cryptic splice acceptor site. Aberrantly spliced transcripts comprise about half of the SPP1 messages expressed by the G allele. Both aberrant splice variants differ from the native transcript by insertions in the leader sequences which do not change the reading frame of SPP1.

CONCLUSIONS

At the 3' terminal end of the first intron of the porcine SPP1 we identified a unique, dually functional SNP g.3836A>G. This SNP affects the function of the SPP1 gene at the DNA level by affecting a C/EBPbeta binding site and at the RNA level by activating aberrant splicing of the first intron, and thus represents an interesting DNA-marker to study phenotypic effects of SPP1 DNA-variation.

Emerging evidence has demonstrated the critical roles for both androgen and Wnt pathways in prostate tumorigenesis. A recent integrative genomic analysis of human prostate cancers (PCas) has revealed a unique enrichment of androgen and Wnt signaling in early-onset PCas, implying their clinical significance in the disease. Additionally, interaction between the androgen receptor (AR) and β-catenin has long been detected in PCa cells. However, the consequence of this interaction in prostate tumorigenesis is still unknown. Because mutations in adenomatous polyposis coli, β-catenin and other components of the destruction complex are generally rare in PCas, other mechanisms of aberrant Wnt signaling activation have been speculated. To address these critical questions, we developed Ctnnb1(L(ex3)/+)/R26hAR(L/+):PB-Cre4 mice, in which transgenic AR and stabilized β-catenin are co-expressed in prostatic epithelial cells. We observed accelerated tumor development, aggressive tumor invasion and a decreased survival rate in Ctnnb1(L(ex3)/+)/R26hAR(L/+):PB-Cre4 compound mice compared with age-matched Ctnnb1(L(ex3)/+):PB-Cre4 littermate controls, which only have stabilized β-catenin expression in the prostate. Castration of the above transgenic mice resulted in significant tumor regression, implying an essential role of androgen signaling in tumor growth and maintenance. Implantation of the prostatic epithelial cells isolated from the transgenic mice regenerated prostate intraepithelial neoplasias and prostatic adenocarcinoma lesions. Microarray analyses of transcriptional profiles showed more robust enrichment of known tumor- and metastasis-promoting genes: Spp1, Egr1, c-Myc, Sp5, and Sp6 genes, in samples isolated from Ctnnb1(L(ex3)/+)/R26hAR(L/+):PB-Cre4 compound mice than those from Ctnnb1(L(ex3)/+):PB-Cre4 and R26hAR(L/+):PB-Cre4 littermate controls. Together, these data demonstrate a confounding role of androgen signaling in β-catenin-initiated oncogenic transformation in prostate tumorigenesis.

OBJECTIVE

This study aims to study how Bio-Oss® can induce osteoblast differentiation in mesenchymal stem cells, the expression levels of bone related genes and mesenchymal stem cells markers using real time Reverse Transcription-Polymerase Chain Reaction.

METHODS

PB-hMSCs stem preparations were obtained for gradient centrifugation from peripheral blood of healthy anonymous volunteers, using the Acuspin System-Histopaque 1077. The samples were then cultured for 7 days for RNA processing, and the expression was quantified using real time PCR.

RESULTS

Bio-Oss® caused an induction of osteoblast transcriptional factor like RUNX2 and of bone related genes; SPP1 and FOSL1. In contrast, the expression of ENG was significantly decreased in stem cells treated with Bio-Oss® with respect to untreated cells, indicating the differentiation effect of this biomaterial on stem cells.

CONCLUSIONS

The results obtained can be relevant to enhance the understanding of the molecular mechanism of bone regeneration and can act as a model for comparing other materials with similar clinical effects.

Physiological occlusal force constitutively exists in the oral environment and is important for periodontal homeostasis and remodeling. Cyclic tensile stress (CTS) triggers the biological response of periodontal ligament (PDL). However, a few reports have studied the correlation between CTS during physiological occlusal force and PDL cell activities such as osteogenic differentiation. In the present study, human PDL cells (hPDLCs) were subjected to 10% elongation CTS loading at 0.5 Hz for 24 h, which represents the physiological conditions of occlusal force. Gene expression microarray was used to investigate the mechano-induced differential gene profile and pathway analysis in vitro. The osteogenic relative factors, that is, SPP1, RUNX2, and SP7, were assessed by real-time PCR and Western blot. The involvement of mitogen-activated protein kinase (MAPK) signaling pathways was investigated by Western blot with a specific inhibitor. The expressions of SPP1, RUNX2, SP7, p-ERK1/2, and p-Elk1 were up-regulated after 10% CTS exposure. However, these up-regulated expressions were prevented by ERK1/2 inhibitor U0126 in the physiological occlusal force-applied hPDLCs. These results showed that 10% CTS could enhance osteogenic differentiation of hPDLCs via ERK1/2-Elk1 MAPK pathway, indicating that CTS during physiological occlusal force is a potent agent for PDL remodeling.

Lung adenocarcinoma (LUAD) is one of the leading causes of cancer death globally. This study aims to investigate the underlying mechanisms implicated with LUAD and identify the key biomarkers. LUAD-associated gene expression dataset (GSE10072) was obtained from GEO database. Based on the GEO2R tool, we screened the differentially expressed genes (DEGs) between the patients with LUAD and normal individuals. Subsequently, Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were employed to find out the enriched pathways of these DEGs. Meanwhile, a protein-protein interaction (PPI) network was also employed to construct to visualize the interactions of these DEGs. Finally, the survival analysis of the top5 up-regulated and top5 down-regulated genes were conducted via GEPIA, aiming to figure out their potential effects on LUAD. In our study, a total of 856 DEGs were captured, including 559 up-regulated genes and 297 down-regulated genes. Among these DEGs, the top5 up-regulated genes were AGER, SFTPC, FABP4, CYP4B1 and WIF1 while the top5 down-regulated genes were GREM1, SPINK1, MMP1, COL11A1 and SPP1. GO analysis disclosed that these DEGs were mainly enriched in DNA synthesis, cell adhesion, signal transduction and cell apoptosis. KEGG analysis unveiled that the enriched pathway included pathways in cancer, PI3K/Akt signaling pathway, MAPK signaling pathway and cell cycle. Survival analysis showed that the expression level of ZG16 may correlate with the prognosis of LUAD patients. Furthermore, according to the connectivity degree of these DEGs, we selected the top15 hub genes, namely IL6, MMP9, EDN1, FOS, CDK1, CDH1, BIRC5, VWF, UBE2C, CDKN3, CDKN2A, CD34, AURKA, CCNB2 and EGR1, which were expected to be promising therapeutic target in LUAD. In conclusion, our study disclosed potential biomarkers and candidate targets in LUAD, which could be helpful to the diagnosis and treatment of LUAD.

Aims. This study is aimed at identifying a prognostic signature for cervical cancer. Main Methods. The gene expression data and clinical information of cervical cancer and normal cervical tissues were acquired from The Cancer Genome Atlas and from three datasets of the Gene Expression Omnibus database. DESeq2 and Limma were employed to screen differentially expressed genes (DEGs). The overlapping DEGs among all datasets were considered the final DEGs. Then, the functional enrichment analysis was performed. Moreover, the Cox proportional hazards regression was performed to establish a prognostic signature of the DEGs. The Kaplan-Meier analysis was applied to test the model. Relationships between gene expression and clinicopathological parameters in cervical cancer, including age, HPV status, histology, stage, and lymph node metastasis, were analysed by the chi-square test. The somatic mutations of these prognostic genes were assessed through cBioPortal. The robustness of the model was verified in another two independent validation cohorts. Key Findings. In total, 169 overlapping upregulated genes and 29 overlapping downregulated genes were identified in cervical cancer compared with normal cervical tissues. Functional enrichment analysis indicated that the DEGs were mainly enriched in DNA replication, the cell cycle, and the p53 signalling pathway. Finally, a 5-gene- (ITM2A, DSG2, SPP1, EFNA1, and MMP1) based prognostic signature was built. According to this model, each patient was given a prognostic-related risk value. The Kaplan-Meier analysis showed that a higher risk was related to worse overall survival in cervical cancer, with an area under the receiver operating characteristic curve of 0.811 for 15 years. The validity of this model in the prediction of cervical cancer outcome was verified in another two independent datasets. In addition, our study also found that the low expression of ITM2A was associated with cervical adenocarcinoma. Interestingly, DSG2 was associated with the HPV status of cervical cancer. Significance. Our study constructed a prognostic model in cervical cancer and discovered two novel genes, ITM2A and DSG2, associated with cervical carcinogenesis and survival.

Cattleyak exhibit equivalent adaptability to harsh environment as yak and much higher performances than yak. However, male infertility of cattleyak due to spermatogenic arrest greatly restricts their effective utilization in yak breeding. Although much work has been done to investigate the mechanisms of spermatogenic arrest, there is little information available in regard to the differences in transcriptomic profiling between cattleyak and yak testes. In this work, histologic observation indicated that spermatogonia were the main type of germ cells present in cattleyak testis, whereas all types of germ cells in differentiation were present in yak testis. Transcriptomic profiling identified 2960 differentially expressed genes (DEGs) in which 679 were upregulated and 2281 were downregulated in cattleyak. Significantly enriched gene ontology terms comprised a large number of DEGs associated with male infertility of cattleyak. The upregulation of STRA8 and NLRP14 may be associated with the accumulation of undifferentiated spermatogonial cells and serious cellular apoptosis in cattleyak. However, downregulated SPP1, SPIN2B, and PIWIL1 were associated with cell cycle progression and spermatogonial genome integrity, whereas CDKN2C, CYP26A1, OVOL1, GGN, MAK, INSL6, RNF212, TSSK1B, TSSK2, and TSSK6 were involved in meiosis. Furthermore, scores of genes associated with sperm components were also downregulated in cattleyak. Wnt/β-catenin signaling pathway was involved in the top-listed three significantly enriched pathways, and the downregulation of Wnt3a, PP2A, and TCF/LEF-1 may have contributed to the arrest of spermatogonial differentiation in cattleyak. The data suggest that spermatogenic arrest of cattleyak might occur at the stage of spermatogonial differentiation and get aggravated during meiosis, which results in minimal number of sperms with morphologic abnormalities and structural deficiency lacking fertilization ability.

The epithelial-to-mesenchymal transition (EMT), which conveys epithelial (E) carcinoma cells to quasi-mesenchymal (qM) states, enables them to metastasize and acquire resistance to certain treatments. Murine tumors composed of qM mammary carcinoma cells assemble an immunosuppressive tumor microenvironment (TME) and develop resistance to anti-CTLA4 immune checkpoint blockade therapy (ICB), unlike their E counterparts. Importantly, minority populations of qM cells within a tumor can cross-protect their more E neighbors from immune attack. The underlying mechanisms of immunosuppression and cross-protection have been unclear. We demonstrate that abrogation of qM carcinoma cell-derived factors (CD73, CSF1 or SPP1) prevents the assembly of an immunosuppressive TME and sensitizes otherwise refractory qM tumors partially or completely to anti-CTLA4 ICB. Most strikingly, mixed tumors in which minority populations of carcinoma cells no longer express CD73, are now sensitized to anti-CTLA4 ICB. Finally, loss of CD73 also enhances the efficacy of anti-CTLA4 ICB during the process of metastatic colonization.

Malignant melanoma is the most deadly form of skin cancer. There is a critical need to identify the patients that could be successfully treated by surgery alone and those that require adjuvant treatment. In this study, we demonstrate that the expression of tribbles2 (TRIB2) strongly correlates with both the presence and progression of melanocyte-derived malignancies. We examined the expression of TRIB2 in addition to 12 previously described melanoma biomarkers across three independent full genome microarray studies. TRIB2 expression was consistently and significantly increased in benign nevi and melanoma, and was highest in samples from patients with metastatic melanoma. The expression profiles for the 12 biomarkers were poorly conserved throughout these studies with only TYR, S100B and SPP1 showing consistently elevated expression in metastatic melanoma versus normal skin. Strikingly we confirmed these findings in 20 freshly obtained primary melanoma tissue samples from metastatic lesions where the expression of these biomarkers were evaluated revealing that TRIB2 expression correlated with disease stage and clinical prognosis. Our results suggest that TRIB2 is a meaningful biomarker reflecting diagnosis and progression of melanoma, as well as predicting clinical response to chemotherapy.

Our previous studies on gastric cancer tissue and patient plasma samples identified several cytokines/chemokines/growth factors to be differentially expressed, compared to normal samples. In this study our aim was to understand the localization patterns of the markers in gastric tissues. We investigated the expression of PDGFRB, CCL3, MMP3, CXCL8, CXCL10, CCL20, IGFBP3, CXCL9, SPP1, CCL18, TIMP1, CCL15, CXCL5 and CCL4 in gastric tissues using Immunohistochemistry (IHC) on Tissue Microarrays (TMA). The TMA comprised of 25 apparently normal (AN), 87 paired normal (PN) and 134 gastric cancer (T) tissues. The epithelial and stromal expression of markers and their correlation with patient characteristics and outcome were analyzed. Several of the markers [PDGFRB (p<0.001), CCL3 (p<0.001), MMP3 (p<0.001), CXCL8 (p<0.001), CXCL10 (p<0.001), CCL20 (p<0.001), CXCL9 (p<0.001), CCL18 (p<0.001), TIMP1 (p=0.025), CCL15 (p<0.001)] were elevated in the stromal compartment of gastric cancers compared to AN tissues, with some having intermediate levels of expression in PN tissues. Epithelial and stromal PDGFRB (p=0.030, p=0.018) expression was associated with diffuse type gastric cancer. Stromal IGFBP3 (p=0.039), CXCL8 (p=0.008), TIMP1 (p<0.001), CCL4 (p=0.003) and SPP1 (p=0.048) expression was associated with intestinal type gastric cancer. Kaplan-Meier analysis showed higher epithelial PDGFRB (p=0.005 and p=0.004), CXCL8 (p=0.009 and p=0.007) were associated with poor disease free and overall survival. In multivariate analysis, high epithelial PDGFRB (p=0.036 and p=0.02) and SPP1 (p=0.003 and p<0.001) were independent prognostic factors for DFS and OS in patients with gastric cancer. The expression of cytokine/chemokine/growth factor markers is higher in the gastric tumor stroma compared to the normal gastric stroma and PDGFRB and SPP1 may serve as potential prognostic factors in gastric cancer.

Follistatin-like protein 1 (FSTL1) plays a critical role in lung organogenesis, but is downregulated during lung cancer development and progression. The prognostic significance and functional consequences of FSTL1 downregulation in lung cancer are unclear. Here, reduced levels of FSTL1 were detected in various tumors compared to normal tissues and were associated with poor clinical outcome in patients with non-small cell lung cancer, particularly those with lung adenocarcinoma. FSTL1 expression negatively correlated with the metastatic potential of lung cancer cells. Antibody-based neutralization of extracellular FSTL1 increased cellular migration/invasion while addition of recombinant FSTL1 protein diminished the metastatic capacity of lung cancer cells in vitro and in vivo. Notably, treatment with FSTL1 effectively prevented the metastatic progression of lung cancer cells in an orthotopic animal model. Mechanistically, FSTL1 directly bound to the pro-form of secreted phosphoprotein 1 (SPP1)/osteopontin restraining proteolytic activation of SPP1 which led to inactivation of integrin/CD44-associated signaling and rearrangement of the actin cytoskeleton. Combined low expression of FSTL1 and high expression of SPP1 predicted a poorer prognosis for patients with lung cancer. This study highlights the novel interaction between FSTL1 and SPP1 and new opportunities to effectively target SPP1-driven metastatic cancers characterized by FSTL1 downregulation.

OBJECTIVE

Recently, genetic pathways that modify the clinical severity of Duchenne muscular dystrophy (DMD) have been identified. The pathways uncovered as modifiers are useful to predict prognosis and also elucidate molecular signatures that can be manipulated therapeutically.

RESULTS

Modifiers have been identified using combinations of transcriptome and genome profiling. Osteopontin, encoded by the SPP1 gene, was found using gene expression profiling. Latent TGFβ binding protein 4, encoding latent TGFβ binding protein 4 was initially discovered using a genome-wide screen in mice and then validated in cohorts of DMD patients. These two pathways converge in that they both regulate TGFβ. A third modifier, Anxa6 that specifies annexin A6, is a calcium binding protein that has been identified using mouse models, and regulates the injury pathway and sarcolemmal resealing.

CONCLUSIONS

Genetic modifiers can serve as biomarkers for outcomes in DMD. Modifiers can alter strength and ambulation in muscular dystrophy, and these same features can be used as endpoints used in clinical trials. Moreover, because genetic modifiers can influence outcomes, these genetic markers should be considered when stratifying results in muscular dystrophy.

We examined whether single nucleotide polymorphisms (SNPs) in SPP1 gene are associated with risk of calcium oxalate urolithiasis (COU). We genotyped nine known SNPs in SPP1 gene (rs11739060, rs28357094, rs2728127, rs11730582, rs1126772, rs9138, rs2853744, rs4754=p.Asp80Asp, and rs1126616=p.Ala236Ala). Genomic DNA from 1,026 individuals (n = 342 patients with first episode COU, and n = 684 healthy unrelated controls) was analyzed for nine SPP1 SNPs using polymerase chain reaction and melting curve analysis by means of a pair of fluorescence resonance energy transfer probes. Serum and urine osteopontin (OPN) levels were also measured using enzyme-linked immunosorbent assay kits. rs9138 AA genotype was protective (OR 0.62, 95 % CI 0.47-0.81; P = 0.004). rs28357094 TT genotype (OR 2.52, 95 % CI 1.74-3.79; P = 0.021), rs2728127 GG genotype (OR 2.64, 95 % CI 1.42-4.81; P = 0.002), and rs2853744 GG genotype (OR 1.68, 95 % CI 1.22-3.87; P = 0.003) were predisposing. None of the other examined SPP1 SNPs was associated with COU susceptibility. Subjects with protective and predisposing polymorphisms had increased and decreased serum levels of OPN, respectively. Urinary calcium/OPN ratios were higher and lower in subjects with predisposing and protective SNPs of SPP1 gene, respectively. Of 28 constructed haplotypes, 6 demonstrated significant association with COU risk. There was no sex difference in the obtained results. The SPP1 gene polymorphisms are associated with the COU susceptibility.

Patients diagnosed with hepatocellular carcinoma (HCC) suffered a high risk of recurrence and poor prognosis. Identification of differentially expressed genes (DEGs) in HCC provides potential biomarkers for evaluating prognosis and specific therapeutic treatments. In this study, DEGs over-expressed in HCC specimens with a fold change over 2.0 were collected through integrative bioinformatics analysis from GEO datasets. Gene ontology and KEGG pathway enrichment were conducted by applying DAVID database. We noticed Secreted phosphoprotein 1 (SPP1) as one of the signature genes up-regulated in HCC tissues with a close relation to the tumor process. Eighty-seven paired HCC specimens from our medical center were explored to verify the aberrant expression of SPP1 by IHC and qRT-PCR assay. Depletion of SPP1 in HCC Hep3B cells was established. The cell proliferation was impaired in SPP1 depleted cells, along with a resistance of cell apoptosis by down-regulating SPP1. Intriguingly, we further validated a direct interaction between miR-181c and SPP1, which indicated a post-transcriptional regulation mechanism of SPP1 in HCC. Thus, our results suggest that SPP1 may function as an enhancer of HCC growth targeted by miR-181c, and probably provide us an innovational target for HCC diagnose and therapeutic treatment.

The cellular distribution of SPP1, which we recently identified as a gene with greater expression in the macaque primary motor cortex than in the premotor or prefrontal cortices, was examined in rhesus macaque, common marmoset, and rat brains. In situ hybridization histochemistry revealed that SPP1 mRNA was expressed specifically in pyramidal neurons in layer V of the sensorimotor cortex of the rhesus macaque. These SPP1 mRNA-positive neurons were most abundant in the primary motor area, followed by Brodmann area 5 and the supplementary motor area, in accordance with the distribution of corticospinal neurons. In addition, injection of a retrograde neuroanatomical tracer into the lateral corticospinal tract (CST) of the spinal cord caused labeling of SPP1 positive neurons, indicating the expression of SPP1 in corticospinal neurons. SPP1 was also expressed in the thalamus, brainstem, and spinal ventral horn of the rhesus macaque. Although SPP1 was also detected in the brainstem and spinal cord of the marmoset and the rat, it was not detected in their cerebral cortices. Selective expression in the corticospinal neurons of the sensorimotor cortex of the rhesus macaque suggests that SPP1 plays a critical role in the functional or structural specialization of highly developed corticospinal systems in certain primate species.

BACKGROUND

Pneumocystis pneumonia is a common opportunistic disease in AIDS patients. The alveolar macrophage is an important effector cell in the clearance of Pneumocystis organisms by phagocytosis. However, both the number and phagocytic activity of alveolar macrophages are decreased in Pneumocystis infected hosts. To understand how Pneumocystis inactivates alveolar macrophages, Affymetrix GeneChip RG-U34A DNA microarrays were used to study the difference in global gene expression in alveolar macrophages from uninfected and Pneumocystis carinii-infected Sprague-Dawley rats.

RESULTS

Analyses of genes that were affected by Pneumocystis infection showed that many functions in the cells were affected. Antigen presentation, cell-mediated immune response, humoral immune response, and inflammatory response were most severely affected, followed by cellular movement, immune cell trafficking, immunological disease, cell-to-cell signaling and interaction, cell death, organ injury and abnormality, cell signaling, infectious disease, small molecular biochemistry, antimicrobial response, and free radical scavenging. Since rats must be immunosuppressed in order to develop Pneumocystis infection, alveolar macrophages from four rats of the same sex and age that were treated with dexamethasone for the entire eight weeks of the study period were also examined. With a filter of false-discovery rate less than 0.1 and fold change greater than 1.5, 200 genes were found to be up-regulated, and 144 genes were down-regulated by dexamethasone treatment. During Pneumocystis pneumonia, 115 genes were found to be up- and 137 were down-regulated with the same filtering criteria. The top ten genes up-regulated by Pneumocystis infection were Cxcl10, Spp1, S100A9, Rsad2, S100A8, Nos2, RT1-Bb, Lcn2, RT1-Db1, and Srgn with fold changes ranging between 12.33 and 5.34; and the top ten down-regulated ones were Lgals1, Psat1, Tbc1d23, Gsta1, Car5b, Xrcc5, Pdlim1, Alcam, Cidea, and Pkib with fold changes ranging between -4.24 and -2.25.

CONCLUSIONS

In order to survive in the host, Pneumocystis organisms change the expression profile of alveolar macrophages. Results of this study revealed that Pneumocystis infection affects many cellular functions leading to reduced number and activity of alveolar macrophages during Pneumocystis pneumonia.

The mechanism of genome transfer from the virion to the host cytoplasm is critical to understand and control the beginning of viral infection. The initial steps of bacteriophage SPP1 infection of the Gram-positive bacterium Bacillus subtilis were monitored by following changes in permeability of the cytoplasmic membrane (CM). SPP1 leads to a distinctively faster CM depolarization than the one caused by podovirus ϕ29 or myovirus SP01 during B. subtilis infection. Depolarization requires interaction of SPP1 infective virion to its receptor protein YueB. The amplitude of depolarization depends on phage input and concentration of YueB at the cell surface. Sub-millimolar concentrations of Ca(2+) are necessary and sufficient for SPP1 reversible binding to the host envelope and thus to trigger depolarization while DNA delivery to the cytoplasm depends on millimolar concentrations of this divalent cation. A model describing the early events of bacteriophage SPP1 infection is presented.