Hepatoprotective effect of prenatal celecoxib in weaning preeclamptic rats: Role of HMGB1/MAPKs signaling.

Preeclampsia (PE) is often associated with multiple organ damage that remains noticeable postnatally. Here, we tested the hypotheses that antenatal therapy with nonsteroidal antiinflammatory drugs (NSAIDs) refashions liver damage induced by PE in weaning rats and that the high mobility group box 1 (HMGB1) signaling modulates this interaction. PE was induced by pharmacologic nitric oxide deprivation during the last week of gestation (Nω-nitro-L-arginine methyl ester, L-NAME, 50 mg/kg/day, oral gavage). Compared with control rats, weaning PE rats revealed substantial rises in serum transaminases together with histopathological signs of hepatic cytoplasmic changes, portal inflammation, and central vein dilation. While gestational NSAIDs reversed the elevated transaminases, they had no effects (celecoxib, naproxen) or even worsened (diclofenac) the structural damage. Molecularly, celecoxib was the most effective NSAID in (i) reversing PE-evoked upregulation of hepatic HMGB1 gene expression and concomitant increments and decrements in mitogen-activated protein kinases MAPKERK and MAPKp38 expression, respectively, and (ii) elevating and suppressing serum interleukin-10 and tumor necrosis factor-α, respectively. Alternatively, rises in serum interleukin-1ß and shifts in macrophage polarization towards an inflammatory phenotype caused by PE were comparably diminished by all NSAIDs. The data disclose an advantageous therapeutic potential for gestational celecoxib over diclofenac or naproxen in controlling hepatic dysfunction and HMGB1-interrelated inflammatory and oxidative sequels of PE.

Modulation by antenatal therapies of cardiovascular and renal programming in male and female offspring of preeclamptic rats.

Morbidity and mortality risks are enhanced in preeclamptic (PE) mothers and their offspring. Here, we asked if sexual dimorphism exists in (i) cardiovascular and renal damage evolved in offspring of PE mothers, and (ii) offspring responsiveness to antenatal therapies. PE was induced by administering NG-nitro-L-arginine methyl ester (L-NAME, 50 mg/kg/day, oral gavage) to pregnant rats for 7 days starting from gestational day 14. Three therapies were co-administered orally with L-NAME, atrasentan (endothelin ETA receptor antagonist), terutroban (thromboxane A2 receptor antagonist, TXA2), or α-methyldopa (α-MD, central sympatholytic drug). Cardiovascular and renal profiles were assessed in 3-month-old offspring. Compared with offspring of non-PE rats, PE offspring exhibited elevated systolic blood pressure and proteinuria and reduced heart rate and creatinine clearance (CrCl). Apart from a greater bradycardia in male offspring, similar PE effects were noted in male and female offspring. While terutroban, atrasentan, or α-MD partially and similarly blunted the PE-evoked changes in CrCl and proteinuria, terutroban was the only drug that virtually abolished PE hypertension. Rises in cardiorenal inflammatory (tumor necrosis factor alpha, TNFα) and oxidative (isoprostane) markers were mostly and equally eliminated by all therapies in the two sexes, except for a greater dampening action of atrasentan, compared with α-MD, on tissue TNFα in female offspring only. Histopathologically, antenatal terutroban or atrasentan was more effective than α-MD in rectifying cardiac structural damage, myofiber separation, and cytoplasmic alterations, in PE offspring. The repair by antenatal terutroban or atrasentan of cardiovascular and renal anomalies in PE offspring is mostly sex-independent and surpasses the protection offered by α-MD, the conventional PE therapy.

Transcriptome profiles of stem-like cells from primary breast cancers allow identification of ITGA7 as a predictive marker of chemotherapy response.

BACKGROUND: Breast cancer stem cells (BCSCs) are drivers of therapy-resistance, therefore are responsible for poor survival. Molecular signatures of BCSCs from primary cancers remain undefined. Here, we identify the consistent transcriptome of primary BCSCs shared across breast cancer subtypes, and we examine the clinical relevance of ITGA7, one of the genes differentially expressed in BCSCs. METHODS: Primary BCSCs were assessed using immunohistochemistry and fluorescently labelled using Aldefluor (n = 17). Transcriptomes of fluorescently sorted BCSCs and matched non-stem cancer cells were determined using RNA-seq (n = 6). ITGA7 expression was examined in breast cancers using immunohistochemistry (n = 305), and its functional role was tested using siRNA in breast cancer cells. RESULTS: Proportions of BCSCs varied from 0 to 9.4%. 38 genes were significantly differentially expressed in BCSCs; genes were enriched for functions in vessel morphogenesis, motility, and metabolism. ITGA7 was found to be significantly downregulated in BCSCs, and low expression significantly correlated with reduced survival in patients treated with chemotherapy, and with chemoresistance in breast cancer cells in vitro. CONCLUSIONS: This study is the first to define the molecular profile of BCSCs from a range of primary breast cancers. ITGA7 acts as a predictive marker for chemotherapy response, in accordance with its downregulation in BCSCs.

Prenatal endothelin or thromboxane receptor antagonism surpasses sympathoinhibition in improving cardiorenal malfunctions in preeclamptic rats.

Current therapies for preeclampsia (PE) and its complications are limited and defective. Considering the importance of endothelin (ET) and thromboxane A2 (TXA2) signaling in PE pathophysiology, we tested the hypothesis that prenatal blockade of endothelin ETA or thromboxane TXA2 receptors favorably reprograms preeclamptic cardiovascular and renal insults. PE was induced by daily oral administration of L-NAME (50 mg/kg) to pregnant rats for 7 consecutive days starting from gestational day 14. The effects of co-exposure to atrasentan (ETA receptor blocker, 10 mg/kg/day) or terutroban (TXA2 receptor blocker, 10 mg/kg/day) on cardiovascular and renal anomalies induced by PE were assessed on gestational day 20 (GD20) and at weaning time and compared with those evoked by the sympatholytic drug α-methyldopa (α-MD, 100 mg/kg/day), a prototypic therapy for PE management. Among all drugs, terutroban was basically the most potent in ameliorating PE-evoked increments in blood pressure and decrements in creatinine clearance. Cardiorenal tissues of PE rats exhibited significant increases in ETA and TXA2 receptor expressions and these effects disappeared after treatment with atrasentan and to a lesser extent by terutroban or α-MD. Atrasentan was also the most effective in reversing the reduced ETB receptor expression in renal tissues of PE rats. Signs of histopathological damage in cardiac and renal tissues of PE rats were mostly improved by all therapies. Together, pharmacologic elimination of ETA or TXA2 receptors offers a relatively better prospect than α-MD in controlling perinatal cardiorenal irregularities sparked by PE.

Peripheral blood and hepatic Toll-like receptor 7 expression and interferon lambda 1 levels in chronic hepatitis C: Relation to virus replication and liver injury.

BACKGROUND AND AIM: Toll-like receptor 7 (TLR7) can recognize single-stranded RNA viruses like hepatitis C virus (HCV) with subsequent induction of different interferon (IFN) types including IFN lambda (IFNL), which activate an immediate anti-viral response. However, the role of TLR7 in inflammation and fibrosis, characteristics of HCV-induced liver injury, is still controversial. The present work was designed to investigate the potential role of TLR7 and IFNL1 in chronic hepatitis C (CHC) in relation to viral replication and liver injury. METHODS: Forty two treatment-naïve patients with CHC and 20 healthy subjects were enrolled in the study. TLR7 expression on peripheral blood CD14+ monocytes was studied by color flow cytometry and the frequency of TLR7+CD14+ cells was expressed as percentage of total monocyte count. Quantification of IFNL1 levels in serum was determined using enzyme-linked immunosorbant assay. Liver biopsies were examined for assessment of histological activity grade (A0-A3) and fibrosis stage (F0-F4) according to METAVIR scoring system as well as steatosis grade. Immunohistochemical staining was performed using human antibodies against TLR7 and IFNL1 and was scored semi-quantitatively (score 0-3). Hepatic expression of TLR7 and IFNL1 was further classified using a two-grade scale as low expression (score 0 or 1) and high expression (score 2 or 3). RESULTS: Percentages of circulating TLR7+CD14+ monocytes and serum IFNL1 levels were significantly higher in patients with CHC than in healthy controls (P = 0.025 and P < 0.001 respectively) and were positively correlated with corresponding hepatic TLR7 and IFNL1 expression (P < 0.001 and P = 0.010 respectively). Significantly lower peripheral blood and hepatic TLR7 expression and IFNL1 levels were found in patients with viral loads between 200,000-600,000 IU/ml and >600,000 IU/ml than in those with viral load <200,000 IU/ml (P < 0.05), in patients with severe necroinflammation than in those with mild-to-moderate necroinflammation (P < 0.05) and in patients with advanced fibrosis than in those with early fibrosis (P < 0.01). Also, changes in TLR7 expression and IFNL1 production in peripheral blood and the liver were inversely correlated with serum levels of aspartate and alanine aminotransferases (P < 0.05) and HCV RNA (P < 0.01), histological activity grade (P < 0.01) and fibrosis stage (P < 0.01). By plotting receiver operating characteristics (ROC) curve, serum IFNL1 showed higher sensitivity and specificity than percentages of circulating TLR7+CD14+ monocytes in discriminating patients with CHC according to the severity of hepatic necroinflammation (area under the curve (AUC) = 0.901 vs. 0.816 respectively) and fibrosis (AUC = 0.971 vs. 0.825 respectively) at a cut-off value of 44.75 pg/ml and 10.25% respectively. CONCLUSIONS: TLR7 activation and IFNL1 production in CHC may play an important role in controlling viral replication and limiting hepatic inflammation and fibrosis and their downregulation may result in viral persistence and disease progression. The immunoregulatory role of TLR7-IFNL1 pathway in the pathogenesis of chronic HCV infection should be further studied. Clinical trials with a large number of patients are needed to assess the usefulness of serum IFNL1 as a potential biomarker for severity of liver injury in chronic HCV infection and other liver diseases.

Modulation by NADPH oxidase of the chronic cardiovascular and autonomic interaction between cyclosporine and NSAIDs in female rats.

Cyclosporine (CSA) and nonsteroidal antiinflammatory drugs (NSAIDs) are used together to manage arthritic disorders with an immune component. Previous reports showed contrasting effects for NSAIDs on CSA nephrotoxicity and acute elevations in blood pressure. Both effects were ameliorated or exaggerated after selective cyclooxygenase-2 (COX2) and nonselective COX inhibition, respectively. Here we investigated: (i) the interaction of CSA with NSAIDs possessing variable COX1/COX2 selectivities on hemodynamic, left ventricular (LV) and cardiac autonomic and histologic profiles, and (ii) role of NADPH-oxidase (NOX)/Rho-kinase (ROCK) pathway in the interaction. Female rats were pre-instrumented with femoral catheters and treated for 10 days with CSA (25mg/kg/day), diclofenac (nonselective NSAIDs, 1mg/kg/day), celecoxib (COX2 inhibitor, 10mg/kg/day), or their combinations. CSA-mediated hypertension was maintained upon co-administration of either NSAID whereas the concomitant reductions in time- and frequency-domain indices of heart rate variability (HRV) were accentuated in presence of diclofenac but not celecoxib. The isovolumic relaxation time (Τau), a measure of diastolic function, was reduced by all regimens whereas LV contractility (dP/dtmax) remained unaffected. The CSA/diclofenac regimen, but not individual treatments, increased cardiac NOX2 expression and caused more cardiac structural damage. The inhibition of NOX by diphenyleneiodonium reversed CSA/diclofenac-evoked increases in MAP, decreases in HRV and Tau, cardiac structural damage, and increased NOX2 expression. No such effects were observed after ROCK inhibition by fasudil, despite concomitant decreases in NOX2 expression. In conclusion, CSA/diclofenac-treated female rats exhibit exacerbated hemodynamic, autonomic, LV, and histopathologic disturbances via ROCK-independent NOX2 upregulation.

Stem cell markers OCT4 and nestin in laryngeal squamous cell carcinoma and their relation to survivin expression.

We explored the expression of the stem cell markers OCT4 and nestin in laryngeal squamous cell carcinoma (LSCC) and investigated their relationship to survivin expression. Eighty-five LSCC, and 62 non-neoplastic laryngeal tissues were analyzed immunohistochemically for the presence of OCT4, nestin and survivin. Marker expression was correlated to clinicopathological parameters. The positive detection rates of OCT4 (42.35%) and nestin (51.76%) in LSCC were higher than those of non-neoplastic mucosa (p<0.05). OCT4 expression was positively associated with nestin expression (p=0.0001). High expression of both OCT4 and nestin was associated with higher tumor grade (p=0.0001). Also, high OCT4 expression was related to higher T stage (p=0.0001). Co-expression of OCT4 and nestin was more significantly associated with glottic location, higher T stage and nodal metastasis than high expression of either marker (p=0.015, 0.006 and 0.008, respectively). Survivin expression was not significantly related to expression of OCT4 or nestin (p=0.094 and 0.266, respectively). OCT4 and nestin are overexpressed in LSCC and may contribute to laryngeal carcinogenesis. Their co-expression may help to predict the lymph node metastatic potential of LSCC. No relationship was detected between expression of survivin and OCT4 or nestin.

Assessment of maturation status of tumor-infiltrating dendritic cells in invasive ductal carcinoma of the breast: relation with vascular endothelial growth factor expression.

OBJECTIVE: Poor immunogenicity has been described in breast carcinoma although dendritic cells, the major antigen presenters, are known to infiltrate the tumor. Vascular endothelial growth factor has been proposed to reduce local immune response in tumors. We investigated the maturation status of dendritic cells in invasive ductal carcinoma of the breast in relation to vascular endothelial growth factor expression and clinicopathological parameters. MATERIAL AND METHOD: Fifty invasive ductal carcinomas of the breast were immunostained with CD1a (marker of immature dendritic cells); CD83 (marker of mature dendritic cells), vascular endothelial growth factor, estrogen receptor and progesterone receptor. RESULTS: Mature dendritic cells were detected in 36 cases (72%), and correlated with smaller tumor size, negative lymph nodes, positive steroid receptor status, and lower grade (P < 0.001). Immature dendritic cells were found in 100% of cases and correlated only with negative steroid receptor expression (estrogen receptor and progesterone receptor) (P=0.006 and 0.020 respectively). Vascular endothelial growth factor expression was detected in 44 cases (88%), and correlated directly with positive nodal metastases (P=0.014), correlated inversely with mature dendritic cell count (P=0.005); and did not correlate with immature dendritic cell count (P=0.104). CONCLUSION: Mature dendritic cell count correlates with good prognostic features in invasive ductal carcinoma of the breast, suggesting their role in initiating primary anti-tumor immune response. Vascular endothelial growth factor expression may play a role in inhibition of dendritic cell maturation sequence in the tumor microenvironment.

Biological and demographic profile of meningiomas in a cohort of Egyptian patients: impact on tumor recurrence.

OBJECTIVE: This work was designed to study the biological and demographic characteristics of meningiomas and their impact on tumor recurrence in Egyptian patients. MATERIAL AND METHODS: A cohort of 265 Egyptian patients with meningioma was studied. Immunohistochemistry for VEGF, Ki67, PR, CD20, and CD3 was performed. Statistical analysis was used to detect independent predictors of recurrence. RESULTS: Adults represented 98.9% of cases, with female preponderance (M : F ratio = 1 : 2.4). Histologically, 78.10% of cases were grade I, 19.20% were grade II, and 2.60% were grade III. Transitional variant was the most common (43.40%). VEGF expression (38.50% of cases) correlated positively with perifocal edema, tumor size, and proliferative index (PI). PR expression (64.5% of cases) correlated inversely with the PI (mean 3.75). Lymphocytic aggregates were detected in 7.20% of cases, with a mean CD20 : CD3 ratio of 1 : 10.1. In a multivariate analysis, only tumor size, PR expression and necrosis predicted recurrence independently. Using ROC curve, size was the best predictor of tumor recurrence with a cut-off point of >6 cm and an excellent negative predictive value (97.6%). CONCLUSIONS: Meningiomas in our region showed some distinctive clinicopathological and demographic criteria. Tumor size was found to be the best recurrence predictor factor of meningioma.