Objective Clinically useful serum biomarkers for the diagnosis and monitoring of eosinophilic gastrointestinal diseases are not available. This study was conducted to examine the possible value of eosinophil-related proteins as serum biomarkers. Methods The serum concentrations of 49 cytokines, chemokines, and other proteins were measured in 29 patients with eosinophilic gastrointestinal diseases and 80 controls. Results The levels of interleukin (IL)-5, IL-33, eotaxin-3, and thymic stromal lymphopoietin (TSLP), previously reported as possible biomarkers of eosinophilic esophagitis, were not significantly elevated in the serum. In contrast, the B cell-attracting chemokine (BCA)-1/chemokine (C-X-C motif) ligand (CXCL) 13 and hemofiltrate C-C chemokine (HCC)-1/CC chemokine ligand (CCL) 14α levels were significantly elevated, while the granulocyte chemotactic protein (GCP)-2/CXCL6 levels were suppressed in patients with eosinophilic esophagitis as well as in those with eosinophilic gastroenteritis. The cutaneus T cell-attracting chemokine (CTACK)/CCL27, stromal cell-derived factor (SDF)-1/CXCL12, macrophage inflammatory protein (MIP)-3β/CCL19, and squamous cell carcinoma antigen (SCCA) 2 levels were elevated only in patients with eosinophilic esophagitis. However, there were large overlaps of data obtained from the patient and control groups, indicating that these serum biomarkers are not adequately sensitive for clinical use with presently available assay systems. Conclusion Of the 49 investigated serum proteins, none were shown to be adequately sensitive for use as biomarkers for the diagnosis or monitoring of eosinophilic gastrointestinal diseases.

Peritoneal dialysis (PD) is a renal replacement option for patients with end-stage renal disease. However, a long-term exposure to hypertonic PD solutions leads to peritoneal membrane (PM) injury, resulting in ultrafiltration (UF) failure. This study was designed to primarily evaluate efficacy of PD effluent-derived mesenchymal stromal cells (pMSCs) in the prevention of PM injury in rats. The pMSCs were isolated from PD effluent. Male Wistar rats received daily intraperitoneal (IP) injection of 10 mL of Dianeal (4.25% dextrose) and were treated with pMSCs (1.2-1.5 × 10⁶/rat/wk, IP). UF was determined by IP injection of 30 mL of Dianeal (4.25% dextrose) with dwell time of 1.5 h, and PM injury was examined by histology. Apoptosis was quantitated by using flow cytometric analysis, and gene expression by using the PCR array and Western blot. Here, we showed that as compared to naive control, daily IP injection of the Dianeal PD solution for 6 weeks without pMSC treatment significantly reduced UF, which was associated with an increase in both PM thickness and blood vessel, while pMSC treatment prevented the UF loss and reduced PM injury and blood vessels. In vitro incubation with pMSC-conditioned medium prevented cell death in cultured human peritoneal mesothelial cells (HPMCs) and downregulated proinflammatory (i.e., CXCL6, NOS2, IL1RN, CCL5, and NR3C1) while upregulated anti-inflammatory (i.e., CCR1, CCR4, IL9, and IL-10) gene expression in activated THP1 cells. In conclusion, pMSCs prevent bioincompatible PD solution-induced PM injury and UF decline, suggesting that infusing back ex vivo-expanded pMSCs intraperitoneally may have therapeutic potential for reduction of UF failure in PD patients.

Gastroduodenitis caused by H. pylori, often acquired in early childhood, is found in about 50% of the adult population. Although H. pylori infections can remain asymptomatic, its virulence factors usually trigger epithelial vacuolization and degeneration, loss of microvilli, disintegration of cytoplasm, and leukocyte accumulation. It is believed that leukocyte infiltration is driven by cytokines produced locally in infected tissue. However, so far little is known about changes in serum cytokines in juvenile patients infected with H. pylori. Serum cytokine profiles were analyzed in 62 juvenile patients diagnosed with gastroduodenitis using the Bio-Plex multiplex assay. H. pylori infection was confirmed in 32 patients, while 30 patients were H. pylori-free. Cytokines CXCL5 and CXCL6, potent neutrophil chemoattractants, were upregulated in all patients diagnosed with gastroduodenitis. Serum levels of IL8, a prototype neutrophil attractant, remained unchanged in subjects with gastroduodenitis relative to controls. Therefore, our data suggest that CXCL5 and CXCL6 play a role in directing neutrophil trafficking into inflamed gastroduodenal tissue. In addition, the CCL25/GM-CSF ratio differed significantly between H. pylori-positive and -negative juveniles. Further, study is needed to evaluate the role of CCL25 and GM-CSF in the pathogenesis of the different etiologies of gastroduodenitis.

Identification of novel circulating biomarkers predicting death and major cardio-metabolic events in obese patients with heart failure (HF) remains a research priority. In this study, we compared multi-marker profile of non-obese (NOB) and obese (OB) HF patients in relation to mortality outcome. The new multiplex proximity extension assay technology was used to analyze the levels of 92 proteins in plasma samples from HF patients according to body mass index (BMI) categories. At 2-year follow-up, all-cause mortality rates were significantly greater in NOB patients (BMI < 30 kg/m2) compared to the OB patients (BMI>> 30 kg/m2) with HF (odds ratio 26; 95% CI: 1.14-624, p < 0,04). Quantitative proteomic analysis revealed thirteen distinct proteins expression profiles of OB and NOB HF patients. Among these proteins, RAGE, CXCL6, CXCL1, CD40, NEMO, VEGF-A, KLK6, PECAM1, PAR1, MMP1, BNP and NTproBNP were down-regulated, whereas leptin was up-regulated in OB HF patients. In addition, an inverse correlation between plasma BNP levels and leptin in OB HF patients was observed (r = -0.58 p = 0.02). This study identifies specific plasma protein signature in OB and NOB patients with HF in relation to mortality outcome.

Interleukin (IL)-25, a member of the IL-17 cytokine superfamily, is produced by immune and non-immune cells and exerts type 2 pro-inflammatory effects in vitro and in vivo. The IL-25 receptor(R) is composed of the IL-17RA/IL-17RB subunits. Previous work showed that germinal centre (GC)-derived B-cell non Hodgkin lymphomas (B-NHL) expressed IL-17AR, formed by IL-17RA and IL-17RC subunits, and IL-17A/IL-17AR axis promoted B-NHL growth by stimulating neoangiogenesis. Here, we have investigated expression and function of IL-25/IL-25R axis in lymph nodes from human GC-derived B-NHL, i.e. Follicular Lymphoma (FL,10 cases), Diffuse Large B Cell Lymphoma (6 cases) and Burkitt Lymphoma (3 cases). Tumor cells expressed IL-25R and IL-25 that was detected also in non-malignant cells by flow cytometry. Immunohistochemical studies confirmed expression of IL-25R and IL-25 in FL cells, and highlighted IL-25 expression in bystander elements of the FL microenvironment. IL-25 i) up-regulated phosphorylation of NFkBp65, STAT-1 and JNK in B-NHL cells; ii) inhibited in vitro proliferation of the latter cells; iii) exerted anti-tumor activity in two in vivo B-NHL models by dampening expression of pro-angiogenic molecules as VEGF-C, CXCL6 and ANGPT3. In conclusion, IL-25, that is intrinsically pro-angiogenic, inhibits B-NHL growth by reprogramming the angiogenic phenotype of B-NHL cells.

Tumor angiogenesis is characterized by deregulated gene expression in endothelial cells (EC). While studies until now have mainly focused on overexpressed genes in tumor endothelium, we here describe the identification of transcripts that are repressed in tumor endothelium and thus have potential suppressive effects on angiogenesis. We identified nineteen putative angiosuppressor genes, one of them being bromodomain containing 7 (BRD7), a gene that has been assigned tumor suppressor properties. BRD7 was studied in more detail, and we demonstrate that BRD7 expression is inversely related to EC activation. Ectopic expression of BRD7 resulted in a dramatic reduction of EC proliferation and viability. Furthermore, overexpression of BRD7 resulted in a bromodomain-dependent induction of NFκB-activity and NFκB-dependent gene expression, including ICAM1, enabling leukocyte-endothelial interactions. In silico functional annotation analysis of genome-wide expression data on BRD7 knockdown and overexpression revealed that the transcriptional signature of low BRD7 expressing cells is associated with increased angiogenesis (a.o. upregulation of angiopoietin-2, VEGF receptor-1 and neuropilin-1), cytokine activity (a.o. upregulation of CXCL1 and CXCL6), and a reduction of immune surveillance (TNF-α, NFκB, ICAM1). Thus, combining in silico and in vitro data reveals multiple pathways of angiosuppressor and anti-tumor activities of BRD7.

Use of porcine tissues has been suggested as a promising solution for severe shortage of transplantable human organs. The immediate hurdle for xenotransplantation is acute immune/inflammatory vascular rejection of the transplant. Because endothelial cells play a key role in the initiation and the amplification of inflammation, alteration of gene expression in human endothelial cells, by various inflammatory stimulators has been studied extensively. However, transcriptional changes induced by human and other inflammatory stimulators in porcine endothelial cells have thus far not been studied. In this study, we treated porcine endothelial cells with human tumor necrosis factor (TNF)-alpha, porcine interferon (IFN)-gamma, H(2)O(2) and lypopolysaccharide (LPS) and profiled transcriptional change at 1 hr, 6 hr and 24 hr, using pig oligonucleotide 13K microarray. We found that mRNA species such as chemokine (C-X-C motif) ligand 6 (CXCL6) and Cathepsin S were significantly induced in porcine endothelial cells, as was previously reported with human endothelial cell. We also found that mRNA species including secreted frizzled-related protein 2 (SFRP2), radical S-adenosyl methionine domain containing 2 (RSAD2), structure specific recognition protein 1 (SSRP1) also were highly overexpressed in porcine endothelial cells. This result shows clues to understand underlying mechanisms of xenotransplantation rejection and the highly responsive porcine genes may serve as novel targets to be regulated for improving the function of grafted porcine donor organs.

The branched-chain fatty acid valproate (valproic acid; VPA) displays antitumoral properties by blocking tumor growth, progression and invasion. Recent data have shown that VPA reduces the angiogenic activity of endothelial cells. The object of this study was to investigate whether endothelial modulation might also influence the level of chemotactic mediators. Endothelial cells were isolated from human umbilical cord veins (HUVEC) and treated with VPA-concentrations ranging from 0.125 mM to 1 mM. The mRNA level of CXC-chemokines was investigated by reverse transcriptase-polymerase chain reaction. The proliferative activity of HUVEC was measured as well. VPA evoked a striking increase in the neutrophil chemoattractants CXCL1, CXCL3, CXCL4, CXCL5 and a moderate increase in CXCL6 with maximal effects after a 3-day incubation period. Other CXC-chemokines and CXC-receptors remained unaffected. HUVEC growth was diminished time- and dose-dependently by VPA. We conclude that VPA treatment leads to alterations in the chemokine expression profile of endothelial cells. This might allow more neutrophils to reach the tumor area and trigger cytolysis.

UNASSIGNED

Rosacea is a chronic facial skin disorder characterized by inflammation and vascular abnormalities. The pathophysiology of rosacea involves increased activation of the capsaicin receptor, TRPV1, the vascular endothelial growth factor (VEGF) pathway, and cathelicidin LL-37, MMP-9, and KLKs. We evaluated the activity of four compounds (dextran sulfate, 4-t-butylcyclohexanol [BCH; TRP-regulin®], pongamia oil, and hesperidin methyl chalcone [HMC]) on inflammatory and vascular responses implicated in rosacea.

UNASSIGNED

The anti-inflammatory activity of dextran sulfate was evaluated on PGE2 production after PMA stimulation of NCTC-2544 keratinocytes, and on normal human epidermal keratinocytes (NHEKs) after proinflammatory stimulation to mimic a rosacea environment. The anti-angiogenic activity of dextran sulfate was measured by analyzing pseudotube formation in co-cultured human microvascular endothelial cells/normal human dermal fibroblasts. HMC modulation of vascular responses and IL-8 cytokine production after SP stimulation was evaluated in human skin explants. We also assessed the effect of BCH on TRPV1 activation, and the effect of combined BCH and pongamia oil on the inflammatory response of NHEKs.

UNASSIGNED

Dextran sulfate strongly and significantly inhibited PMA-induced PGE2 production, inhibited KLK5 and MMP-9 mRNA expression, and IL-8, IL-1α and VEGF production, and displayed a highly significant inhibitory effect on VEGF-induced pseudotube formation. In SP-stimulated human skin explants, HMC significantly decreased the proportion of dilated vessels, total vessel area, and IL-8 production. BCH significantly and dose-dependently inhibited TRPV1 activation, and BCH and pongamia oil inhibited CXCL1 and CXCL6 mRNA expression and IL-8 production in NHEKs. Combined BCH/pongamia oil inhibited IL-8 production synergistically.

UNASSIGNED

These in vitro results showed that dextran sulfate, BCH, pongamia oil and HMC, possess complementary soothing and anti-redness properties, supporting their combination in Avène redness-relief cosmetic products for sensitive skin prone to redness, and for topical adjunctive rosacea treatment.

BACKGROUND

In renal cell carcinoma (RCC) of the clear cell type, inactivity of the VHL gene induces overexpression of HIF1 α and its targets, the tyrosine kinase receptors, promoting RCC development and progression. The discovery of tyrosine kinase inhibitors (TKIs) changed the treatment of these tumors. Other molecular pathways involved in the TKI mechanisms of action have not been described in the literature. The aim of our study was to elucidate alternative mechanisms of action of sunitinib in tumor tissue after neoadjuvant treatment of RCC.

METHODS

The gene expression profile was accessed using microarray (Affymetrix Human Genome U133 Plus 2.0 platform) and frozen RCC tissues collected from 5 patients with locally advanced non-metastatic tumors who underwent nephrectomy after being treated with 2 cycles of neoadjuvant sunitinib. The results were compared with matched controls comprising 6 patients with no neoadjuvant intervention.

RESULTS

There was underexpression of the majority of genes after sunitinib treatment. The lower expression levels of IGFBP1, CCL20, CXCL6 and FGB were confirmed by qRT-PCR in all cases. The downregulation of gene expression leads us to search for methylation as a mechanism of action of the TKI. IGFBP1 was shown to be methylated by methylation-sensitive high-resolution melting technique.

CONCLUSIONS

The ultimate genetic effects of sunitinib may explain its actions as an antitumor drug that apparently suppresses the expression of important genes related to cell survival, adhesion, invasion and immunomodulation. The methylation of gene promoters was shown to be part of the mechanism of action of this class of drugs.

BACKGROUND

The objective of this study was to characterize the mRNA expression profile related to rumen epithelial inflammation through the in vivo and in vitro experiments. In the in vivo experiment, rumen papillae were collected from four dairy cows adapted to either a 40 % (LC) or 70 % (HC) concentrate feeds for microarray analysis.

RESULTS

Results showed that 245 differentially expressed genes (DEGs) were detected in the cows fed the HC relative to the LC diet. The DEGs were first annotated, and results revealed that the expression of inflammation-related genes, including IL-1β, IL-2, IL-22, CCL19, CCL8, CX3CR1, CXCL6, INHBE, LEPR, PRL, and TNFRSF9 found in the cytokine-cytokine receptor pathway were up-regulated in the HC-fed cows, indicating local inflammation in the rumen epithelium was triggered. The expression of IL-1β, IL-2, and IL-6 was further validated by qRT-PCR. To demonstrate whether there were relationships between cytokine mRNA expression and ruminal factors (pH and LPS), the isolated ruminal epithelial cells were cultured in vitro. Results showed that the mRNA expression of IL-1β, IL-2, IL-6, and IL-8 increased after the LPS treatment, while low-pH treatment elevated the mRNA expression of TNF-α, suggesting that low-pH coupled with higher levels of LPS in rumen of cows fed the HC may be mainly responsible for the triggered local ruminal inflammation.

CONCLUSIONS

Our results indicate that ruminal local inflammation response might be triggered during HC feeding, and these findings also enhance the knowledge of rumen epithelial adaptation to HC at the molecular level.

The key event in placentitis-induced preterm labor is myometrial activation with the subsequent initiation of labor. However, the molecular mechanisms underlying myometrial activation are not fully understood in the mare. Therefore, the equine myometrial transcriptome was characterized during placentitis (290.0 ± 1.52 d GA, n = 5) and the prepartum period (330d GA, n = 3) in comparison to normal pregnant mares (289.8 ± 2.18 GA, n = 4). Transcriptome analysis identified 596 and 290 DEGs in the myometrium during placentitis and the prepartum period, respectively, with 138 DEGs in common. The placentitis DEGs included eight genes (MMP1, MMP8, S100A9, S100A8, PI3, APOBEC3Z1B, RETN and CXCL2) that are exclusively expressed in the inflamed myometrium. Pathway analysis elucidated that inflammatory signaling, toll-like receptor signaling, and apoptosis pathways dominate myometrial activation during placentitis. Prepartum myometrium was associated with overexpression of inflammatory signaling, oxidative stress, and 5-hydroxytryptamine degradation. Gene ontology enrichment analysis identified several chemoattractant factors in the myometrium during placentitis and prepartum period, including CCL2, CXCL1, CXCL3, and CXCL6 in common. Upstream regulator analysis revealed 19 potential upstream regulators in placentitis dataset including transcription regulators (E2F1, FOXM1, HIF1A, JUNB, NFKB1A, and STAT1), transmembrane receptors (FAS, ICAM1, SELP, TLR2, and TYROBP), growth factors (HGF and TGFB3), enzymes (PTGS2 and PRKCP), and others (S100A8, S100A9, CD44 and C5AR1). Additionally, three upstream regulators (STAT3, EGR1, and F2R) were identified in the prepartum dataset. These findings revealed the key regulators and pathways underlying myometrial activation during placentitis, which aid in understanding the disease and facilitate the development of efficacious therapies.

Bromodomain-containing protein 4 (BRD4) has been implicated to play a regulatory role in fibrogenic gene expression in animal models of liver fibrosis. The potential role of BRD4 in liver fibrosis in humans remains unclear. We sought to investigate the expression and cellular localization of BRD4 in fibrotic liver tissues. Human liver tissues were collected from healthy individuals and patients with liver fibrosis of various etiologies. RNA-seq showed that hepatic BRD4 mRNA was elevated in patients with liver fibrosis compared with that in healthy controls. Subsequent multiple manipulations such as western blotting, real-time quantitative polymerase chain reaction, and dual immunofluorescence analysis confirmed the abnormal elevation of the BRD4 expression in liver fibrosis of various etiologies compared to healthy controls. BRD4 expression was positively correlated with the severity of liver fibrosis, and also correlated with the serum levels of aspartate aminotransferase and total bilirubin. Moreover, the expression of C-X-C motif chemokine ligand 6 (CXCL6), a factor interplayed with BRD4, was increased in hepatic tissues of the patients with liver fibrosis. Its expression level was positively correlated with BRD4 level. BRD4 is up-regulated in liver fibrosis, regardless of etiology, and its increased expression is positively correlated with higher degrees of liver fibrosis. Our data indicate that BRD4 play a critical role in the progress of liver fibrosis, and it holds promise as a potential target for intervention of liver fibrosis.

Keywords: BRD4; CXCL6; HBV; human samples; liver fibrosis.

A functional genomics approach was used to examine the immune response for transcriptional profiling of PBMC M. bovis infected cattle and healthy control cattle to stimulation with bovine tuberculin (purified protein derivative PPD-b). Total cellular RNA was extracted from non-challenged control and M. bovis challenged MDM for all animals at intervals of 6 h post-challenge, in response to in-vitro challenge with M. bovis (multiplicity of infection 2:1) and prepared for global gene expression analysis using the Agilent Bovine (V2) Gene Expression Microarray, 8 × 60 K. The pattern of expression of these genes in PPD bovine stimulated PBMC provides the first description of an M. bovis specific signature of infection that may provide insights into the molecular basis of the host response to infection. Analysis of these mapped reads showed 2450 genes (1291 up regulated and 1158 down regulated) 462 putative natural antisense transcripts (354 up-regulated and 108 down regulated) that were differentially expressed based on sense and antisense strand data, respectively (adjusted P-value ≤ 0.05). The results provided enrichment for genes involved top ten up regulated and down regulated panel of genes, including transcription factors proliferation of T and B lymphocytes. The highest differentially-expressed genes were associated to immune and inflammatory responses, immunity, differentiation, cell growth, apoptosis, cellular trafficking and regulation of lipolysis and thermogenesis. Microarray results were confirmed in infected cattle by RT qPCR to identify potential biomarkers TLR2, CD80, NFKB1, IL8, CXCL6 and ADORA3 of bovine tuberculosis.

Diabetic nephropathy (DN) is one of the main causes of renal fibrosis and is associated with high morbidity and mortality. Traditional Chinese Medicine (TCM) therapy has a long history of usage in a clinical setting and its usage is increasing. ErHuang Formula (EHF), a Chinese herbal compound, has been clinically used in treating DN for more than 30 years. However, its mechanism of action is still unknown. This study was conducted to evaluate the effect of EHF on renal fibrosis in a DN rat model and explore its underlying mechanism. The DN rat model was established by high-sugar-fat diet combined with a single intraperitoneal injection of streptozotocin (STZ), and EFH extract (4, 2, 1 g/kg d^-1) was administered orally for 8 weeks. The biochemical parameters (blood glucose, weight, Scr, BUN, UA, U-Alb and UAE) were analyzed. The pathological changes in renal tissue were observed by histological staining with H&E and Masson. The effect of EHF on the proliferation of NRK-49F cells was examined by CCK-8 assay and the levels of several inflammation and fibrosis related cytokines (IL-6, TNF-α, TGF-β1, Collagen I/III, MMP2/9) in serum and NRK-49F cell culture supernatants were detected by enzyme-linked immunoassay (ELISA). The mRNA levels of CXCL6, CXCR1, Collagen I/III, MMP2/9 in renal tissue were also measured by quantitative RT-PCR. Furthermore, the protein expression of PCNA, Collagen I/III, MMP2/9, CXCL6, CXCR1, p-STAT3, STAT3 in renal tissue and NRK-49F cells were determined by western blot. EHF improved the abnormal biochemical parameters and ameliorated the abnormal histology and fibrosis of renal tissue in a dose-dependent manner. EHF inhibited NRK-49F proliferation and decreased the expressions of inflammation and fibrosis related factors both in vitro and in vivo. Interestingly, the levels of Collagen I/III, PCNA, MMP2/9 and p-STAT3 were positively correlated with CXCL6. The amelioration of renal fibrosis in DN by EHF is related to CXCL6/JAK/STAT3 signal pathway, which is associated with inflammation and fibrosis of the tissue. These findings may have clinical implications for the treatment of DN.

Primary IgA nephropathy (IgAN) diagnosis is based on IgA-dominant glomerular deposits and histological scoring is done on formalin-fixed paraffin embedded tissue (FFPE) sections using the Oxford classification. Our aim was to use this underexploited resource to extract RNA and identify genes that characterize active (endocapillary-extracapillary proliferations) and chronic (tubulo-interstitial) renal lesions in total renal cortex. RNA was extracted from archival FFPE renal biopsies of 52 IgAN patients, 22 non-IgAN and normal renal tissue of 7 kidney living donors (KLD) as controls. Genome-wide gene expression profiles were obtained and biomarker identification was carried out comparing gene expression signatures a subset of IgAN patients with active (N = 8), and chronic (N = 12) renal lesions versus non-IgAN and KLD. Bioinformatic analysis identified transcripts for active (DEFA4, TNFAIP6, FAR2) and chronic (LTB, CXCL6, ITGAX) renal lesions that were validated by RT-PCR and IHC. Finally, two of them (TNFAIP6 for active and CXCL6 for chronic) were confirmed in the urine of an independent cohort of IgAN patients compared with non-IgAN patients and controls. We have integrated transcriptomics with histomorphological scores, identified specific gene expression changes using the invaluable repository of archival renal biopsies and discovered two urinary biomarkers that may be used for specific clinical decision making.

Mesenchymal stromal cells derived from the fetal placenta, composed of an amnion membrane, chorion membrane, and umbilical cord, have emerged as promising sources for regenerative medicine. Here, we used next-generation sequencing technology to comprehensively compare amniotic stromal cells (ASCs) with chorionic stromal cells (CSCs) at the molecular and signaling levels. Principal component analysis showed a clear dichotomy of gene expression profiles between ASCs and CSCs. Unsupervised hierarchical clustering confirmed that the biological repeats of ASCs and CSCs were able to respectively group together. Supervised analysis identified differentially expressed genes, such as LMO3, HOXA11, and HOXA13, and differentially expressed isoforms, such as CXCL6 and HGF. Gene Ontology (GO) analysis showed that the GO terms of the extracellular matrix, angiogenesis, and cell adhesion were significantly enriched in CSCs. We further explored the factors associated with inflammation and angiogenesis using a multiplex assay. In comparison with ASCs, CSCs secreted higher levels of angiogenic factors, including angiogenin, VEGFA, HGF, and bFGF. The results of a tube formation assay proved that CSCs exhibited a strong angiogenic function. However, ASCs secreted two-fold more of an anti-inflammatory factor, TSG-6, than CSCs. In conclusion, our study demonstrated the differential gene expression patterns between ASCs and CSCs. CSCs have superior angiogenic potential, whereas ASCs exhibit increased anti-inflammatory properties.

Keywords: RNA-seq; amniotic stromal cells; angiogenesis; anti-inflammation; chorionic stromal cells; placenta.

Bacillus Calmette-Guérin (BCG) immunotherapy is the standard therapy for non-muscle invasive bladder cancer. The aim of the present study was to identify genes that are induced in response to BCG immunotherapy, as these may be potential biomarkers for the response to clinical therapy. To model clinical therapy, human bladder cancer cell lines were incubated with BCG (live or lyophilized BCG Connaught) for 2 h. RNA was extracted and evaluated by Representational Differential Analysis (RDA) and oligo arrays. Gene expression was confirmed by reverse transcription polymerase chain reaction on fresh cell lines with differential abilities to internalize BCG. The effect of 2 major BCG soluble proteins, antigen 85B (Ag85B) and Mycobacterium protein tyrosine phosphatase A (MptpA) and BCG Tice® on gene expression was also determined. GAPDH and β-actin, which are normally used as control genes, were upregulated by BCG. Therefore, the ribosomal RNA gene ribosomal protein S27a was used to normalize gene expression. The genes likely to be induced by BCG internalization and soluble factors were: GSTT2, MGST2, CCL20, TNFα, CCNE1 and IL10RB. Those induced by BCG membrane interactions and/or soluble factors were: MGST1, CXCL6, IL12A, CSF2, IL1β and TOLLIP. MptpA decreased GSTT2 expression, and Ag85B increased TNFα expression. The two BCG strains significantly increased GSTT2, TNFα and TOLLIP levels in MGH cells. However, in J82 cells there was a BCG strain-dependent difference in TNFα expression. An important outcome of the present study was the determination that neither GAPDH nor β-actin were suitable control genes for the analysis of BCG-induced gene expression. BCG Connaught and Tice® induced similar expression levels of genes in bladder cancer cell lines. BCG soluble proteins modulated gene expression and therefore may affect therapeutic outcomes. The genes identified may be novel biomarkers of the response to BCG therapy.

Abnormalities of neuroinflammation have been implicated in the pathogenesis of depression and suicide. This is primarily based on the observation that cytokines, which are major inflammatory molecules and play an important role in depression and suicide, are increased in both serum and in postmortem brain of depressed and suicidal subjects. Another class of immune mediators are chemokines which are primarily involved in chemotactic properties and trafficking of immune cells in the central nervous system (CNS). Chemokines also play an important role in CNS function. Whereas chemokines have been studied in the serum of depressed and suicidal patients, their role in brain of depressed or suicidal subjects is relatively unexplored. We studied the gene expression of several chemokines in the prefrontal cortex (PFC) obtained from depressed suicidal (DS) and normal control (NC) subjects. We determined the mRNA expression of several chemokines belonging to CXCL and CCL groups of chemokines using qPCR array technique and qPCR gene expression validation in 24 DS and 24 NC subjects. The postmortem brain samples were obtained from the Maryland Brain Collection. We found that the mRNA expression of chemokines CXCL1, CXCL2, CXCL3 and CCL2 was significantly decreased in the PFC of DS compared with NC subjects. No significant change was observed in CXCL5, CXCL6, CXCL10, CCL8 and CCL19 between DS and NC subjects. Since many of the chemokines are involved in mediating certain important CNS functions, such as neurotrophic effect, neurogenesis, anti-apoptotic growth factor release, modulation of synaptic transmission, brain development and neuronal loss, decreased levels of chemokines can reduce these functions which may be involved in the pathophysiology of depression.

Keywords: chemokines; depression; postmortem brain; prefrontal cortex (PFC), CCL2, CXCL1; suicide.

The Caspase activation and recruitment domain 8 (CARD8) protein is a component of innate immunity and overexpression of CARD8 mRNA was previously identified in atherosclerosis. However, very little is known about the regulation of CARD8 in endothelial cells and atherosclerosis. The aim of this study was to investigate CARD8 in the regulation of cytokine and chemokine expression in endothelial cells. Sections of human atherosclerotic lesions and non-atherosclerotic arteries were immunostained for CARD8 protein. Expression of CARD8 was correlated to mediators of inflammation in atherosclerotic lesions using Biobank of Karolinska Endarterectomies microarray data. The CARD8 mRNA was knocked-down in human umbilical vein endothelial cells (HUVECs) in vitro, followed by quantitative RT-PCR analysis and OLINK Proteomics. Endothelial and smooth muscle cells in arterial tissue expressed CARD8 and CARD8 correlated with vWF, CD163 and the expression of inflammatory genes, such as CXCL1, CXCL6 and PDGF-A in plaque. Knock-down of CARD8 in HUVECs significantly altered proteins involved in inflammatory response, such as CXCL1, CXCL6, PDGF-A, MCP-1 and IL-6. The present study suggest that CARD8 regulate the expression of cytokines and chemokines in endothelial cells and atherosclerotic lesions, suggesting that CARD8 plays a significant role in endothelial activation.

In subacute rumen acidosis (SARA), the rumen epithelium is frequently attacked by endotoxin (LPS), which is caused by the lysis of dead Gram-negative bacteria. However, the rumen epithelium innate immune system can actively respond to the infection. Previous studies have demonstrated that tea tree oil (TTO) has good bactericidal and anti-inflammatory effects. Therefore, the aim of this study was to investigate the effect of TTO on the expression of genes involved in the inflammatory cytokines in goat rumen epithelial cells (GRECs) triggered by LPS. Our study shows that rumen epithelial cells isolated from goat rumen tissue can be cultured in vitro in 0.25% trypsin for a long time. These cells were identified as epithelial cells by the expression of cytokeratin 18, monocarboxylate transporter 4 (MCT4), Na[+]/H[+] hydrogen exchanger 1 (NHE1), putative anion transporter 1 (PAT1), vH⁺ ATPase B subunit (vH⁺ ATPase), and anion exchanger 2 (AE2). The mRNA expression of IL-1β, IL-6, TNF-α, TLR-2, NF-κB, CXCL6 and CXCL8 genes was significantly increased when LPS was used compared to untreated controls. In addition, mRNA expression of IL-1β, IL-6, TNF-α, TLR-2, NF-κB, CXCL8, CXCL6 and interferon-induced protein with tetratricopeptide repeats 3 (IFIT3) genes was also significantly higher in the LPS group compared to the 0.05% TTO group. However, the expression of IL-1β, IL-6, TNF-α, TLR-2, CXCL6 and IFIT3 genes was significantly lower in the LPS and 0.05% TTO group compared to the 1 μg/mL LPS group. These results suggest that TTO can inhibit LPS-induced inflammatory cytokines expression in GRECs.

Keywords: LPS; goat rumen epithelial cells; innate immune; tea tree oil.

Breast cancer (BC) in adolescents and young adults (AYAs) accounts for 5.6% of BC in all women. BC in this population is often characterized as aggressive. Two-thirds of BC in AYAs belong to the hormone-receptor positive (HR(+))/human epidermal growth factor receptor 2 negative (HER2(-)) subtype. However, the underlying pathogenesis of this subtype has not been fully elucidated. To study HR(+)/HER2(-) BC in AYAs, we downloaded the available RNA-seq data from The Cancer Genome Atlas (TCGA) database and then performed differential expression gene screening and constructed a protein-protein interaction (PPI) network, identified the key genes, and did gene set enrichment analysis (GSEA). Based on the analyses, 32.26% of patients were in stage III. Additionally, we identified 1671 differentially expressed genes (DEGs) and 35 key genes. In addition, GSEA showed that ether lipid metabolism and complement and coagulation cascades were significantly enriched in the GNAI1 high expression phenotype. The key genes CXCL2, CXCL5, CXCL3, GPR37L1, NPY2R, OXGR1, NPW, CCL21, GNAI1, SAA1, GRM4, HCAR2, CX3CL1, GRM8, CCL28, SSTR1, PENK, P2RY12, NMUR1, NMU, ADCY5, TAS1R1, OXER1, GNG13, CCL16, CCR8, NPY5R, CXCL11, CXCL10, CXCL9, CXCL1, CXCL6, CCR4, and ANXA1 may be molecular markers of tumorigenesis of HR(+)/HER2(-) BC in AYAs. In addition, ether lipid metabolism and complement and coagulation cascades may be key pathways for GNAI1 regulation in HR(+)/HER2(-) BC in AYAs.

Keywords: Breast cancer; HR(+)/HER2(-); adolescents and young adults; differential expression; pathway analysis.

Objectives: To investigate the distribution of multiple cytokines in gastroschisis and reveal its association with clinical outcomes, including gastrointestinal disorders and fetal brain damage caused by chronic inflammation in gastroschisis.

Methods: We obtained amniotic fluid and arterial cord blood from 10 patients with gastroschisis, and evaluated the profile of 40 cytokines via multiplex immunoassay. The possible relationship of the cytokines with the time taken to attain full enteral nutrition and cord S100B, a surrogate marker of brain damage, was estimated. Associations among the relevant cytokines were also assessed.

Results: Although clinical characteristics in our cohort had no relevance, several cytokines in cord blood, especially IL-2, IL-8, CCL1, CCL7, CXCL1, CXCL2, and CXCL6, were clearly elevated in patients who took a longer time to attain full enteral nutrition, whereas only IL-16 in cord blood was significantly related to cord S100B and strongly correlation with cord S100B levels. Moreover, our data indicated that IL-16 was considerably less correlated with the other cytokines associated with adverse outcomes.

Conclusions: We investigated the cytokine characteristics of both amniotic fluid and cord blood in gastroschisis, and found that certain cytokines could affect the adverse outcomes, including fetal brain damage. These findings provide important information that could further clarify the pathophysiology of gastroschisis and propose a novel clinical implication of gastroschisis that could be used to predict adverse outcomes, especially neurodevelopmental disorders.

Keywords: Amniotic fluid; Biomarker; Pregnancy; Umbilical cord.