Effect of prolactin on cytotoxicity and oxidative stress in ovine ovarian granulosa cells.

Background: Prolactin (PRL) has been reported to be associated with oxidative stress, which is an important contributor leading to cell apoptosis. However, little is known about the mechanisms underlying the effects of PRL on cytotoxicity and oxidative stress in ovine ovarian granulosa cells (GCs). Methods: Ovine ovarian GCs were treated with 0, 4, 20, 100 and 500 ng/mL of PRL. Then, the cytotoxicity, cell viability, malondialdehyde (MDA), reactive oxygen species (ROS), superoxide dismutase (SOD) and total antioxidant capacity (T-AOC) of GCs were detected. Additionally, 500 ng/mL PRL was chosen as the high PRL concentration (HPC) due to its high cytotoxicity and oxidative stress. Proteomic and metabonomic were performed to examine the overall difference in proteins and metabolic pathways between C (control: 0 ng/mL PRL) and P groups (500 ng/mL PRL). Results: The results indicated that GCs treated with 4 ng/mL PRL significantly decreased (P < 0.05) the cytotoxicity, ROS and MDA, increased (P < 0.05) the cell viability, SOD and T-AOC, and the GCs treated with 500 ng/mL PRL showed the opposite trend (P < 0.05). Supplementation with 500 ng/mL PRL significantly increased the proteins of MT-ND1, MAPK12, UBA52 and BCL2L1, which were enriched in ROS and mitophagy pathways. Pathway enrichment analysis showed that the pentose phosphate pathway was significantly enriched in the P group. Conclusion: A low concentration of PRL inhibited cytotoxicity and oxidative stress. HPC induced oxidative stress in ovine ovarian GCs via the pentose phosphate pathway by modulating the associated proteins MT-ND1 in ROS pathway and UBA52, MAPK12 and BCL2L1 in mitophagy pathway, resulting in cytotoxicity.

Dose analysis of photobiomodulation therapy on osteoblast, osteoclast, and osteocyte.

The objective of this study was to evaluate the effects of varying light doses on the viability and cellular activity of osteoblasts, osteocytes, and osteoclasts. A light application device was developed to apply 940-nm wavelength light from light-emitting diodes on three cultured cells, MC3T3-E1, MLO-A5, and RANKL-treated RAW264.7 cells. The doses (energy density) on cells were 0, 1, 5, and 7.5 J / cm2. The corresponding light power densities at the cell site were 0, 1.67, 8.33, and 12.5 mW / cm2, respectively, and the duration was 10 min. The results showed that the three cell types respond differently to light and their responses were dose dependent. Low-dose treatment (1 J / cm2) enhanced osteoblast proliferation, osteoclast differentiation, and osteoclastic bone resorption activity. Osteocyte proliferation was not affected by both low- and high-dose (5 J / cm2) treatments. While 1 J / cm2 did not affect viability of all three cell types, 5 J / cm2 significantly decreased viability of osteocytes and osteoclasts. Osteoblast viability was negatively impacted by the higher dose (7.5 J / cm2). The findings suggest that optimal doses exist for osteoblast and osteoclast, which can stimulate cell activities, and there is a safe dose range for each type of cell tested.

Changes in nitric oxide, angiotensin â ¡ angiopoietin-like protein 4 mRNA, neuregulin 1 mRNA, and platelet endothelial cell adhesion molecule-1 in rats with acute blood stasis induced by high-molecular-weight dextran.

OBJECTIVE: To investigate the influence of acute blood stasis on nitric oxide (NO), angiotensin Ⅱ(AngⅡ), angiopoietin-like protein 4 (ANGPTL4) mRNA, neuregulin 1 (NRG-1) mRNA, and platelet endothelial cell adhesion molecule-1 (PECAM-1) in rats with stasis induced by high-molecular-weight dextran (HMWD). METHODS: Seventy-five Sprague Dawley rats were divided randomly into five groups (n = 15 in each group): control group, immediate group, 1 h group, 3 h group, and 6 h group. A model of acute blood stasis was established via injection of HMWD into the tail vein. After performing electrocardiogram at the predetermined times according to the grouping, we collected blood and cardiac samples for hematoxylin-eosin (HE) staining and histopathological examination via transmission electron microscopy. Enzyme-linked immunosorbent assay was used to detect plasma levels of NO, AngⅡ, and fibrinogen. Real-time polymerase chain reaction was used to detect the expression of ANGPTL4 mRNA and NRG-1 mRNA. Immunohistochemical methods were used to detect PECAM-1 protein expression. RESULTS: The rat model of blood stasis showed blood retention in the capillary lumens. The ST segment showed gradual elevation, and was still elevated at 3 and 6 h after induction of blood stasis. HE staining showed myocardial cell necrosis and dissolution after modeling, along with basement membrane rupture and mitochondrial structural damage. Transmission electron microscopy showed endothelial cell swelling and an increase in absorption vesicles immediately after modeling. Endothelial cell apoptosis was increased at 3 and 6 h after modeling. Cardiac muscle fibers were disordered and intercalated discs were blurred immediately after modeling. There were massive numbers of dissolved cardiac muscle fibers, ruptured basement membranes, and mitochondrial structural damage at 3 and 6 h after modeling. NO plasma concentration was significantly reduced immediately and 1 h after modeling, while it was increased at 3 and 6 h. Ang¢ò plasma concentration was decreased immediately after modeling, but increased at 1, 3, and 6 h. Fibrinogen plasma concentration was significantly increased at immediate, 1, 3, and 6 h after modeling. PECAM-1 protein expression was obviously increased immediately after modeling, at 1, 6 h was found mild augment. Expression of AngPTL4 mRNA was increased at immediate, 1, 3, and 6 h after modeling, and was found further augment at 3, and 6 h. Expression of NRG-1 mRNA was increased at immediate, 1, 3, and 6 h after modeling, and the strongest expression was at 1 h. CONCLUSION: The pathological manifestation of acute blood stasis is characterized by microvascular blood retention. Prolonged blood stasis leads to worsening endothelial cell and cardiomyocyte damage, along with imbalances in the expression of vasomotor factors and increased vascular tone. The pathological damage caused by blood stasis also promotes the expression of cell protection factors.

Microvascular pathological features and changes in related injury factors in a rat acute blood stasis model.

OBJECTIVE: To examine the microvascular pathological characteristics and changes in related injury factors in a rat model of acute blood stasis. METHODS: A total of 75 Sprague-Dawley rats were divided randomly and equally into a control group and four experimental groups assessed at different times after the induction of stasis (0, 1, 3 or 6 h after stasis) (n = 15). The acute blood stasis model was established through rat tail-vein injection of high-molecular-weight dextran. After Electrocardiograph (ECG) detection at predetermined times (0, 1, 3 and 6 h after induction of stasis), the rats were sacrificed and blood and cardiac samples were harvested for analysis. Hematoxylin-eosin (HE) staining and transmission electron microscopy were used for histopathological detection; an enzyme linked immunosorbent assay (ELISA) was used to detect thromboxane B2 (TXB2) and 6-keto-prostaglandin F1α (6-Keto-PGF1α) concentrations; a real-time polymerase chain reaction (PCR) reaction system was used to detect intercellular adhesion molecule 1 (ICAM-1) and vascular cell adhesion molecule1 (VCAM-1) mRNA expression; western blotting was used to detect vascular endothelial cadherin (VE-cadherin) protein expression. RESULTS: The ST segment in the ECG showed gradual elevation after induction of stasis and continued elevation at a high level at 3 and 6 h. The HE staining showed changes in myocardial cell necrosis and tissue dissociation after the induction of stasis, along with inflammatory infiltration. Results of transmission electron microscopy showed immediate changes in blood stasis and lumen occlusion in the microvasculature, along with endothelial cell swelling. After the induction of stasis, TXB2 concentrations gradually increased while 6-Keto-PGF(1α) concentrations were immediately significantly reduced. The TXB(2)/6-Keto-PGF(1α) ratio was maintained at a high level. ICAM-1 mRNA expression showed an unstable elevation while VCAM-1 mRNA expression was significantly reduced after the induction of stasis. Compared with the control group, VE-cadherin protein expression increased at 0 and 3 h after the induction of stasis, while no change occurred at 1 and 6 h. CONCLUSION: The pathological manifestations of acute blood stasis are microvascular blood retention, lumen stenosis and even occlusion. The condition is also called "blood coagulation and weep" in Traditional Chinese Medicine. The blood stasis model resulted in the injury and necrosis of endothelial cells and cardiomyocytes, along with the presence of an imbalance of vasomotor factor levels, platelet activation, and increases in the expression of adhesion molecules and endothelial barrier dysfunction, which corresponds to "blood failed to nourish" in Traditional Chinese Medicine.

Prereferral head and neck cancer treatment: compliance with national comprehensive cancer network treatment guidelines.

OBJECTIVE: to evaluate the prereferral treatment of patients referred to our tertiary care center with recurrent or persistent head and neck cancer for compliance with National Comprehensive Cancer Network (NCCN) guidelines. DESIGN: a prospective recruitment and retrospective chart review. PATIENTS: the study included new patients identified at multidisciplinary treatment planning conference from October 1, 2008, to February 1, 2009, who had received prior treatment at an outside institution and presented to our department with recurrent or persistent disease. MAIN OUTCOME MEASURES: all facets of prior care were examined, including the time from initial symptoms to diagnosis and whether their prereferral treatment was compliant with or deviated from NCCN guidelines for head and neck cancer. RESULTS: a total of 566 consecutive new patients were identified, of whom 107 (18.9%) had persistent or recurrent disease. The average time from first presentation with initial symptoms to diagnosis among patients who presented with persistent disease was 23.8 weeks. Nearly half of the patients who presented with persistent or recurrent disease had either endocrine (21.5%) or cutaneous (24.2%) primary cancers, with the rest of the cases being distributed among 10 other sites. Of the patients who presented with recurrent or persistent disease, 43.0% had prereferral care that was noncompliant with NCCN guidelines. Of these patients, 58.7% had inadequate surgical management, 15.2% were treated for the wrong diagnosis, 10.9% received inadequate adjuvant therapy, 4.4% received inadequate radiotherapy, and 10.9% refused indicated recommended treatment. CONCLUSIONS: significant deviation from NCCN guidelines for head and neck cancer treatment was observed in the cohort of study patients. The failure to administer adjuvant therapy when indicated by NCCN guidelines is particularly concerning. Economic and noneconomic costs, including lost wages, cost of "do-over" therapy, and potentially diminished survival, are substantial. Measures to ensure that patients receive therapy according to guidelines should be a national priority.