Beta mRNA Research Articles

Tumor expression of human chorionic gonadotropin beta mRNA and prognosis of prostate cancer treated by radical prostatectomy

The beta subunit of human chorionic gonadotropin (hCGβ) is encoded by six genes (CGB) classified as type I and type II. CGB mRNA is produced in large amounts by trophoblastic tissues and in small amounts by several cancerous tissues including prostate cancer and by a few benign tissues, including the prostate. Quantitative reverse-transcription polymerase chain reaction (RT-qPCR) was used to study the expression levels of all CGB mRNAs together (total CGB mRNA) and the two types of CGB mRNA separately in non-cancerous (n = 74) and cancerous prostatic tissue obtained by radical prostatectomy (n = 193). RNA was isolated from formalin-fixed paraffin-embedded (FFPE) samples and mRNA levels of CGB were correlated with disease-specific survival. Total CGB mRNA concentrations were significantly lower (p < .0001) in cancerous than non-cancerous prostatic tissue. Separate analysis of type I CGB and type II CGB mRNA showed that both type I CGB (p < .0001) and type II CGB mRNA (p = .007) are lower in cancerous tissue than in non-cancerous tissue. Low type II CGB mRNA level in cancerous tissue was associated with shorter cancer-specific survival (p = .001) of prostate cancer patients treated by radical prostatectomy.

2182-P: Protective Effects of Lin28a-RNA Binding Protein Against Glucotoxicity in Pancreatic Beta Cell

In pancreatic beta cell, excessive and prolonged increase of glucose causes beta cell dysfunction which is characterized by impaired insulin synthesis and secretion, termed the glucotoxicity. Although it is known that both Lin28a and LIN28B promoted an insulin-sensitized state that resisted high fat diet-induced diabetes, other molecular mechanisms that protect from glucotoxicity are poorly defined. We investigated the effect of Lin28a overexpression on glucotoxicity-induced beta cell dysfunction. In this study, we found that high glucose leads to a decrease in Lin28a mRNA and protein expression, followed by a decrease in insulin gene expression and secretion in the rat insulinoma cell line, INS-1. In addition, PDX-1and BETA2 mRNA and protein expression were downregulated by high glucose in INS-1. Similarly, mRNA expression of Lin28a and insulin was decreased in pancreas of high fat diet induced diabetic mouse. Adenovirus mediated Lin28a overexpression in INS-1 reversed the glucotoxicity-induced downregulation of insulin, PDX-1 and BETA2 expression, and improved glucose stimulated insulin secretion (GSIS). In mouse islet, mRNA expressions of insulin, PDX-1 and ki67 and GSIS were decreased by glucotoxicity. Lin 28a overexpression in mouse islet partially restored their expression and GSIS. These results suggest that Lin28a protects pancreatic beta cells from glucotoxicity through regulation of insulin transcription factors. Disclosure K. Lee: None. Y. Hwang: None. J. Choi: None. Funding Daegu Gyeongbuk Institute of Science and Technology (18-LC-01, 2018010133)

Role of GnRH in the ontogeny and regulation of the fetal hypothalamo-pituitary-gonadal axis in sheep

Adult reproductive ability is to a large extent determined by the appropriate development of the reproductive axis during fetal life. Studies have investigated the role of the fetal hypothalamus in the ontogeny and regulation of pituitary gonadal function during fetal development in sheep. Using immunocytochemistry, we examined the ontogeny of gonadotroph development in the pituitary of female sheep fetuses. At day 70 of gestation (term = 145 days), only immunopositive LH beta cells were present. The number and intensity of staining of these LH beta cells had increased by day 100 but had declined again by day 130. Immunopositive alpha-subunit and FSH beta cells appeared at day 100 of gestation and had further increased in number and staining intensity by day 130 of gestation. Treatment of fetuses with the GnRH agonist buserelin resulted in desensitization of the fetal pituitary gonadotrophs, inhibition of pituitary LH beta and FSH beta mRNA expression and a reduction in the number of immunopositive gonadotrophin-containing cells. Pulsatile GnRH treatment resulted in pituitary-gonadal activation and an increase in LH, FSH and testosterone secretion in males. Thus, the synthesis and secretion of the gonadotrophins during fetal development is critically dependent on the secretion of GnRH from the fetal hypothalamus. Inhibition of fetal gonadotrophins by buserelin treatment from day 70 of gestation resulted in a 40% reduction in the size of the fetal testis at birth, and there were no effects on the fetal ovaries. This reduction in testis size was due to a 45% reduction in the number of Sertoli cells. However, when buserelin was given between day 70 and day 110 of gestation, there were no effects on testis size or morphological development of the testis, suggesting that gonadotrophins regulate testicular development during a 'critical window' late in gestation. Taken together, these studies provide convincing evidence that GnRH plays a central role in the ontogeny and regulation of pituitary-gonadal function during fetal life.

Correlation between luteinizing hormone receptor gene expression in human granulosa cells with oocyte quality in poor responder patients undergoing in vitro fertilization: A cross-sectional study

Background: This study was performed to evaluate the role of luteinizing hormone (LH) and granulosa cell LH receptor (LH-R) in poor responder patients who underwent controlled ovarian stimulation. Expression levels of LH-R mRNA in granulosa cells was investigated and compared with oocyte morphology, oocyte maturity and fertilization rate. Methods: Granulosa cells were obtained from 30 patients who underwent in vitro fertilization (IVF) at Dr. Cipto Mangunkusumo Hospital, Jakarta. The patients were divided into two groups: group I (n=10) poor responders; and group II (n=20) non-poor responders. After the extraction of total RNA from granulosa cells, semi-quantitative RT-PCR was performed and the amount of LH-R mRNA was quantified. The relative values were calculated as the ratio of LH-R mRNA and actin beta mRNA. Statistical analysis was performed using Mann-Whitney test and Spearman correlation. Results: The relative value of LH-R mRNA was higher in group I compared with group II (27.37[0.00-28939.37] vs 0.00[0.00-7196.12]). Oocyte maturity (r=0.267) and morphology (r=0.267) in group I consistently showed a positive correlation with LH-R mRNA; in group II a negative correlation with LH-R mRNA was shown for oocyte maturity (r= -0.552) and morphology (r= -0.164). Group I had a positive correlation between LH-R expression with fertilization rate (r=0.430), and group II showed a negative correlation (r=-0.340). Conclusions: The expression of LH-R mRNA has a positive correlation with oocyte quality in poor responder patients and a negative correlation in non-poor responders. Our study suggests an optimal expression of LH- R mRNA in granulosa cells during controlled ovarian stimulation to obtain good quality oocytes.

Effect and molecular regulatory mechanism of monochromatic light colors on the egg-laying performance of Yangzhou geese

Photoperiodic control is essential for manipulating the reproductive performance of avian species. This study was conducted to assess the neuroendocrine mechanisms that regulate reproductive functions of Yangzhou geese when there are different monochromatic light colors from light emitter diode (LED) sources. A flock of geese was divided into four groups with white, red, blue, and green light treatments being imposed. The results indicated that peak laying rates and reproductive performance were greater in geese treated with white or red as compared with blue or green light treatments. The fertilization rate of eggs and hatchability of fertilized eggs were greater with the white or red as compared with blue or green light treatments. There was a greater abundance of OPN5, Dio2, c-Fos, and GnRH-I mRNA in the hypothalamus earlier in the treatment period and abundances of these hypothalamic factors were greater with the white or red light treatments. Abundances of pituitary LH beta and FSH beta mRNA increased at a lesser rate with the blue or green light treatments and were in greater abundances with the white or red light treatments. The lighting regimen also resulted in photo-refractoriness with there being greater abundances of GnIH, VIP, and PRL mRNA with the use of white or red light treatments. The results indicate that the use of white or red monochromatic lights while imposing a long photoperiod of 11 h daily could result in sustaining functions of the reproductive system of Yangzhou geese for considerably longer times, thus, resulting in greater egg-laying performance.

Identification of metabolic pathways related to the bisphenol A-induced adipogenesis in differentiated murine adipocytes by using RNA-sequencing

We evaluated the effect of bisphenol A and its metabolites on the 3T3-L1 cells, in terms of glucose and lipid metabolism. We also aimed to obtain the information on the genome-wide expression changes in the 3T3-L1 cells treated with Bisphenol A by using RNA-seq, which involves whole-transcriptome sequencing. Differentially Expressed Genes (DEGs) collected from RNA-seq can be used to produce a complete picture of related metabolism pathways. The KEGG pathway was extracted based on the DEGs. Bisphenol A significantly increased the mRNA level of Sterol regulatory element binding transcription factor 1 (Srebf1) and CCAAT/enhancer binding protein alpha (Cebpa). Lipoprotein lipase (Lpl) was also significantly influenced by bisphenol A and its metabolites. Acetyl-Coenzyme A carboxylase beta (Acacb) and Fatty acid synthase (Fasn) mRNA levels were elevated by bisphenol A and its metabolites. The insulin signaling pathway, neurotrophin signaling pathway, and endometrial cancer-related pathway were focused by the functional enrichment analyses, and the pathways were well coincided with recent previous reports. DEGs collected from RNA-seq were confirmed as a reliable evidence in the exposure to the chemicals such as bisphenol A. Collecting pieces of the puzzles obtained from the RNA-seq will help us to produce a complete picture of the metabolic pathway for such chemicals.

Correlation between luteinizing hormone receptor gene expression in human granulosa cells with oocyte quality in poor responder patients undergoing in vitro fertilization: A cross-sectional study.

Background: This study was performed to evaluate the role of luteinizing hormone (LH) and granulosa cell LH receptor (LH-R) in poor responder patients who underwent controlled ovarian stimulation. Expression levels of LH-R mRNA in granulosa cells was investigated and compared with oocyte morphology, oocyte maturity and fertilization rate. Methods: Granulosa cells were obtained from 30 patients who underwent in vitro fertilization (IVF) at Dr. Cipto Mangunkusumo Hospital, Jakarta. The patients were divided into two groups: group I (n=10) poor responders; and group II (n=20) non-poor responders. After the extraction of total RNA from granulosa cells, semi-quantitative RT-PCR was performed and the amount of LH-R mRNA was quantified. The relative values were calculated as the ratio of LH-R mRNA and actin beta mRNA. Statistical analysis was performed using Mann-Whitney test and Spearman correlation. Results: The relative value of LH-R mRNA was higher in group I compared with group II (27.37[0.00-28939.37] vs 0.00[0.00-7196.12]). Oocyte maturity (r=0.267) and morphology (r=0.267) in group I consistently showed a positive correlation with LH-R mRNA; in group II a negative correlation with LH-R mRNA was shown for oocyte maturity (r= -0.552) and morphology (r= -0.164). Group I had a positive correlation between LH-R expression with fertilization rate (r=0.430), and group II showed a negative correlation (r=-0.340). Conclusions: The expression of LH-R mRNA has a positive correlation with oocyte quality in poor responder patients and a negative correlation in non-poor responders. Our study suggests an optimal expression of LH- R mRNA in granulosa cells during controlled ovarian stimulation to obtain good quality oocytes.

Effects of plane of nutrition and arginine on ovarian follicles in non-pregnant sheep: Cell proliferation, and expression of endothelial nitric oxide and its receptor

The aim of this study was to investigate the role of the nitric oxide (NO) system in ovarian function, by determining if arginine (Arg) supplementation impacts follicle number, cell proliferation, and expression of the NO system members in nutritionally compromised ewes. Ewes were randomly assigned into maintenance (C, 100% requirements), excess (O; 2xC), or restricted (U; 0.6xC) diets 8 weeks prior to Arg treatment. Ewes were individually fed twice daily with pelleted diets. Ewes from each nutritional group were randomly assigned to one of two treatments: saline or Arg, which was initiated on day 0 of the estrous cycle and administered 3 times per day. Ovaries were collected at the early-luteal, mid-luteal and late-luteal/follicular phases of the estrous cycle to determine 1) the number of surface follicles, 2) follicle cell proliferation marked by Ki67 protein expression, and 3) expression of endothelial nitric oxide (eNOS; NOS3) and soluble guanylyl cyclase beta (sGC; GUCY1B3) protein and mRNA in granulosa (G) and theca (T) layers using immunohistochemistry followed by image analysis and qPCR, respectively. During nutritional treatment, C maintained body weight, O gained 6±1.2 kg, and U lost 14±1.3 kg. Our data show that: 1) Ki67 was expressed in all ovarian compartments, eNOS protein was detected in blood vessels of T and stroma, and sGC protein was detected in T cells, and blood vessels of T layer and other ovarian compartments; 2) plane of nutrition affected the number of surface follicles, and thus folliculogenesis, cell proliferation in the T layer, eNOS and sGC protein expression in T, and NOS3 and GUCY1B3 mRNA expression in G; 3) Arg treatment affected cell proliferation in G and T, eNOS and sGC protein expression in T, mRNA expression of NOS3 in T in all groups, and GUCY1B3 in G depending on the stage of the estrous cycle; and 4) G and T cell proliferation, and expression of eNOS and sGC protein in T was affected by the stage of the estrous cycle. Our data demonstrated that plane of nutrition and Arg are involved in the regulation of follicular functions in non-pregnant sheep.

Role of endothelin‑1 and its receptors in cerebral vasospasm following subarachnoid hemorrhage.

Cerebral vasospasm (CVS) is a severe complication of subarachnoid hemorrhage (SAH), and endothelin‑1 (ET‑1) may be involved in its pathogenesis. The present study aimed to investigate the expression of ET‑1 in cerebrospinal fluid (CSF) in patients with SAH and to analyze rat arterial contractility and the expression levels of ET‑1 receptors invitro. CSF samples were collected from 28patients and the expression levels of ET‑1 were measured. Rat cerebral basilar arteries were isolated and incubated with hemorrhagic or clear CSF. Contractility, as well as ETA and ETB mRNA expression were measured. ET‑1 levels in CSF increased and reached a peak within the initial 5days after SAH onset and then gradually subsided. After 12 or 24h, the contraction of arteries incubated in hemorrhagic CSF was substantially stronger than those in clear CSF. The mRNA expression levels of endothelin receptor type A and B in arteries incubated in hemorrhagic CSF were significantly higher than those in clear CSF. ET‑1 and its receptors may be involved in the pathogenic mechanism of CVS following SAH. ET‑1 expression in CSF may be used as a marker in CVS and its receptors may provide novel therapeutic targets in CVS.

Detection and localization of the thyroid hormone receptor beta mRNA in the immature olfactory receptor neurons of chum salmon

Thyroid hormone (TH) plays an important role in regulating multiple cellular and metabolic processes, including cell proliferation, cell death, and energy metabolism, in various organs and tissues of vertebrates. It is generally accepted that anadromous Pacific salmon (Oncorhynchus spp.) imprint odorants from their natal stream during their seaward migration, and they then use olfaction to discriminate their natal stream during the spawning migration. Both serum TH levels and the specific binding values of TH in the salmon olfactory epithelium were markedly increased during the seaward migration. However, thyroid hormone receptor (TR) expression in the olfactory epithelium has not been confirmed in vertebrates. We investigated gene expression of TR isoforms in chum salmon (O. keta) by both molecular biological and histochemical techniques. Expression of TRβ mRNA was detected in the olfactory epithelium by reverse transcriptase polymerase chain reaction (RT-PCR). Nucleotide sequencing demonstrated the existence of a remarkable homology between the RT-PCR product and part of the ligand-binding domain of other teleost TRβ isoforms. By in situ hybridization using a digoxygenin-labeled salmon olfactory TRβ cRNA probe, signals for salmon olfactory TRβ mRNA were observed preferentially in the perinuclear regions of immature olfactory receptor neurons (ORNs), as protein gene product 9.5 (PGP9.5)-immunopositive ORNs. Our results provide the first detection of TRβ gene expression in the olfactory epithelium, and suggested the possibility that TRβ may be involved in cell maturation and/or cell differentiation of the ORNs in Pacific salmon.

Conditioned medium from stimulated macrophages inhibits growth but induces an inflammatory phenotype in breast cancer cells

Disparate roles exist for tumor-associated macrophages in breast cancer growth and progression. The aim of this study was to explore the influence of induced macrophages on the growth of breast cancer cells. THP-1 monocytes were differentiated to macrophages using phorbol 12-myristate 13-acetate. The effect of the medium from THP-1 monocytes or macrophage-conditioned medium (MφCM) on MCF-7 (estrogen receptor and progesterone-positive positive) and MDA-MB-231 (MB; triple-negative) breast cancer cells was determined at 24 h, 48 h and 72 h. Assays were conducted for cell viability, apoptosis, proliferation and cell phenotype, and quantitative real-time polymerase chain reaction (qRT-PCR) for expression of associated genes. MφCM inhibited proliferation of MCF-7 and MB cells in a time-dependent manner and, in particular, decreased viability of MCF-7 cells. MφCM induced a markedly vacuolated phenotype in MCF-7 increased apoptosis in MCF-7 cells, but correlative changes in Bcl-2 or Bax were absent. A multifold and significant reduction in anti-apoptotic Bcl-2 in MB cells was not matched by increased apoptosis. The cell cycle inhibitor CDKN1A was increased in both cell lines, but PCNA decreased only in MB cells. Senescence-associated galactosidase beta-1 (GLB1) mRNA was decreased in MCF-7 cells (48 and 72 h) but increased in MB cells (72 h). Increased expression of interleukin-6 (IL-6) and IL-8 was seen in both cell lines, and increased tumor necrosis factor- α was seen at 24 h for MB and 72 h for MCF-7 indicating increased inflammatory responses of the cancer cells. The two breast cancer celllines had different responses to MφCM, mainly involving inhibition rather than stimulation of growth of the cells, stimulation of senescence (MB cells) and increased inflammatory cytokine expression. The estrogen and progesterone receptor status of the cell lines may determine their response to MφCM. The function of the inflammatory cytokines in breast cancer growth remains to be identified.

Nephron prorenin receptor deficiency alters renal medullary endothelin-1 and endothelin receptor expression.

The endothelin (ET) and prorenin/renin/prorenin receptor (PRR) systems have opposing physiological effects on collecting duct (CD) salt and water reabsorption. It is unknown if the CD ET and renin/PRR systems interact, hence we examined the effects of deleting CD renin or nephron PRR on CD ET system components. PRR knockout (KO) mice were polyuric and had markedly increased urinary ET-1 and inner medullary CD (IMCD) ET-1 mRNA. PRR KO mice had greatly increased IMCD ETA receptor mRNA and protein, while ETB mRNA and protein were decreased. Water loaded wild-type mice with similar polyuria as PRR KO mice had modestly increased urinary ET-1 excretion and inner medullary ET-1 mRNA, while inner medullary ETA and ETB mRNA or protein expression were unaffected. In contrast to PRR KO, CD prorenin/renin KO did not alter ET system components. Taken together, these results suggest that the nephron PRR is involved in regulating CD ET system expression, but this effect may be independent of CD-derived renin.

The glucagon-like peptide-1 receptor agonist liraglutide improves hypoxia-induced pulmonary hypertension in mice partly via normalization of reduced ET(B) receptor expression.

The glucagon-like peptide-1 receptor (GLP-1R) agonist liraglutide is an incretin hormone mimetic used in the treatment of diabetes. However, the effects of liraglutide on pulmonary hypertension (PH) and pulmonary endothelin (ET) system are unknown. Eight-week-old C57BL6/J mice were injected liraglutide or vehicle for 5 weeks. One week after injection, the mice were exposed to either room air (normoxia) or chronic hypoxia (10 % O(2)) for 4 weeks. The right ventricular systolic pressure (RVSP) was significantly higher in hypoxia + vehicle group than in normoxia + vehicle group. ET-1 mRNA expression in the lungs was comparable among all the groups. ET(B) mRNA and protein expression in the lungs was significantly lower in hypoxia + vehicle group than in normoxia + vehicle group. The above changes were normalized by liraglutide treatment. The expression of phospho-eNOS and phospho-AMPK proteins in the lungs was significantly higher in hypoxia + liraglutide group than in normoxia + vehicle group. We demonstrated for the first time that liraglutide effectively improved RVSP and RV hypertrophy in hypoxia-induced PH mice by activating eNOS through normalization of impaired ET(B) pathway and augmentation of AMPK pathway. Therefore, GLP-1R agonists can be promising therapeutic agents for PH.

Anti-fibrotic effects of polygonum plebeium r.br. in CCl4-induced hepatic damage and fibrosis in rats

Introduction: Asian countries have the highest burden of liver diseases. Polygonum plebeium (P. plebeium), a.k.a. the common knotweed, is a species of plant in the knotwood family that can act as a blood purifier and has been widely used in Pakistan to cure liver disorders like jaundice and hepatitis. The plant is also used in the treatment of pneumonia, bowel complaints, diarrhea, dysentery, eczema and ring worms. Tannins, flavonoids, saponins and alkaloids are the major components of P. plebeium. Since its use in folk medicine in Pakistan, there has been little scientific evidence or information on it. Therefore, this study was aimed at investigating the anti-fibrotic effects of P. plebeium in carbon tetrachloride (CCl4)-induced hepatic toxicity and fibrosis.&#x0D; Methods: The extracts of whole plant of P. plebeium were prepared and administered by oral gavage in rats. Liver fibrosis was induced by intraperitoneal (i.p.) administration of CCl4. To evaluate the hepatoprotective activity of P. plebeium, alanine aminotransferase (ALT), aspartate aminotransferase (AST), and gamma-glutamyl transpeptidase (γGT) levels were quantified. Histological evaluation of liver tissue revealed tissue necrosis and extracellular matrix deposition. Real-time PCR was done to evaluate mRNA expression of genes related to liver fibrosis.&#x0D; Results: The groups treated with P. plebeium extract showed an ablation in liver damage; the elevated enzyme levels of ALT, AST and γGT were decreased. Treatment with P. plebeium extract treatment restored the CCl4-induced tissue fibrosis by significantly suppressing alpha-smooth muscle actin (α-SMA), tumor growth factor beta (TGF-β) and collagen mRNA expression levels. Histology of liver sections also showed that the CCl4-induced fibrosis was improved in the treatment groups.&#x0D; Conclusion: Polygonum plebeium has therapeutic potential and can be used for preventing fibrosis in inflammatory liver disease.

Nest box exploration may stimulate breeding physiology and alter mRNA expression in the medial preoptic area of female European starlings.

Environmental resources are proposed to fine-tune the timing of breeding, yet how they may do so remains unclear. In female European starlings (Sturnus vulgaris), nest cavities are limited resources that are necessary for breeding. Females that explore nest cavities, compared with those that do not, readily perform sexually motivated behaviors. We assigned female starlings to aviaries with: (1) no nest boxes, (2) nest boxes, or (3) nest boxes, plants, flowing water, insects and berries to test the hypothesis that environmental resources alter neural systems to stimulate mating behavior. Compared with other females, females that were housed with and explored nest boxes had higher estradiol, higher preproenkephalin (PENK) mRNA and lower levels of D1 and D2 dopamine receptor mRNA in the medial preoptic area (mPOA); a region in which opioids and dopamine modify female sexual behaviors and sexual motivation. Additionally, in the mPOA, PENK and tyrosine hydroxylase mRNA positively predicted, whereas estrogen receptor beta mRNA negatively predicted, nest box exploration. In the ventromedial hypothalamus (a region in which estradiol acts to stimulate sexual behavior), estrogen receptor alpha mRNA was highest in females that had access to but did not explore nest cavities. It is likely that seasonal increases in estradiol modify mRNA in the mPOA to facilitate nest cavity exploration. It is also possible that nest cavity exploration further alters gene expression in the mPOA, functioning to coordinate mating with resource availability. Thus, nest cavity exploration may be a form of self-stimulation that alters neural systems to fine-tune sexual behavior.

Interplay of foot and mouth disease virus with cell-mediated and humoral immunity of host.

Foot and mouth disease virus (FMDV) causes a communicable disease of cloven hoofed animals, resulting in major economic losses during disease outbreaks. Like other members of the Picornaviridae FMDV has a relatively short infectious cycle; initiation of infection and dissemination, with production of infectious virions occurs in less than a week. The components of innate immunity as well as cell-mediated and humoral immunity play a crucial role in control of FMDV. However, it has been shown in vitro using a mouse model that FMDV has evolved certain mechanisms to counteract host immune responses ensuring its survival and spread. The viral leader proteinase, L pro, deters interferon beta (IFN-β) mRNA synthesis, thus, inhibiting host cell translation. Another viral proteinase, 3C pro, disrupts host cell transcription by cleaving histone H3. A transient lymphopenia in swine as a consequence of FMDV infection has also been observed, but the mechanism involved and viral protein(s) associated with this process are not clearly understood. In this review, we have covered the interaction of FMDV with different immune cells including lymphocytes and antigen presenting cells and their consequences.

Inflammation and oxidative stress in corneal tissue in experimental keratitis due to Fusarium solani: Amelioration following topical therapy with voriconazole and epigallocatechin gallate.

Combined antifungal and antioxidant therapy may help to reduce oxidative stress in fungal keratitis. Experimental Fusarium solani keratitis was induced by application of F.solani conidia to scarified cornea (right eye) of 16 rabbits (another four rabbits were negative controls [Group I]). Five days later, F.solani-infected animals began receiving hourly topical saline alone (Group II), voriconazole (10mg/mL) alone (Group III), epigallocatechin gallate (EGCG, 10mg/mL) alone (Group IV) or voriconazole and EGCG (Group V). Twenty days post-inoculation, corneal lesions were graded. After animal sacrifice, excised corneas underwent histopathological and microbiological investigations. Corneal tissue levels/activities of interleukin 1 beta (IL-1β) and tumour necrosis factor alpha (TNF-α) gene mRNA transcripts, matrix metalloproteinase (MMP) 2 and 9 proteins, malondialdehyde (MDA) and reduced glutathione (GSH), and superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx), were also measured. Clinical and histopathological scores (severity of corneal lesions; [P<.05]) and mean levels (P<.05) of IL-1β and TNF-α mRNA transcripts, MMP 2, MMP 9 and MDA were Group II>Groups IV and III>Groups V and I. Mean SOD, CAT, GPx and GSH levels (P<.05) were Group II<Groups IV and III<Groups V and I. Topical voriconazole with EGCG apparently reduces inflammation in experimental F.solani keratitis, as manifested by improved clinical, histological, microbiological and molecular parameters.

Estrogen-dependent modifications to hippocampal plasticity in paternal California mice (Peromyscus californicus)

In many biparental species, mothers and fathers experience similar modifications to circulating hormones. With these modifications come alterations in neural structure and function suggesting that neuroendocrine mechanisms may underlie postpartum plasticity in both males and females. In the biparental California mouse (Peromyscus californicus), adult neurogenesis is maintained and anxiety-like behavior is attenuated in fathers during the mid-postpartum period. Given a causal relationship between estrogen and regulation of both adult neurogenesis and anxiety, we aimed to elucidate the role of estrogen-dependent mechanisms in paternal experience-related modifications to hippocampal neuroplasticity in California mice. In Experiment 1, hippocampal estrogen receptor beta (ERβ) mRNA expression, along with circulating estradiol concentrations, were determined throughout the postpartum period. An upregulation in ERβ expression was observed in postnatal day 16 males compared to virgins. Additionally, a rise in circulating estradiol concentrations was detected on postnatal day 2 compared to virgins; levels began to decline toward virgin levels on postnatal day 16 and postnatal day 30. In Experiment 2, we determined the role of estrogen-dependent mechanisms in adult neurogenesis and anxiety-like behavior by treating virgin and paternal males with saline or the selective estrogen receptor modulator, tamoxifen (TMX), during the time of axon extension (i.e., one week after bromodeoxyuridine injection). While TMX failed to alter elevated plus maze performance, TMX treatment inhibited survival of adult born neurons but only in paternal mice. These findings highlight the potential for estrogen-dependent pathways to mediate hippocampal adult neurogenesis in paternal mice.

Potential therapeutic impact of omega-3 long chain-polyunsaturated fatty acids on inflammation markers in Duchenne muscular dystrophy: A double-blind, controlled randomized trial

Duchenne Muscular Dystrophy (DMD) is the most frequent dystrophy in childhood generated by a deficiency in dystrophin. DMD is a neuromuscular disease and its clinical course comprises chronic inflammation and gradual muscle weakness. Supplementation of omega-3 long chain-Polyunsaturated Fatty Acids (ω-3 long chain-PUFA) reduces inflammatory markers in various disorders. The goal of this research was to analyze the influence of ω-3 long chain-PUFA intake on gene expression and blood inflammatory markers in boys with DMD. In a placebo-controlled, double. Blind, randomized trial, boys with DMD (n=36) consumed 2.9g/day of ω-3 long chain-PUFA or sunflower oil as control, in capsules, for a period of 6 months. Blood was analyzed at baseline and at months 1, 2, 3, and 6 of supplementation for expression of inflammatory markers in leukocytes and serum. There was high adherence to capsule intake (control: 95.3%±7.2%, and ω-3 long chain-PUFA: 97.4%±3.7% at month 6). Enrichment of EicosaPentaenoic Acid (EPA) and DocosaHexaenoic Acid (DHA) in erythrocytes increased significantly in patients supplemented with ω-3 long chain-PUFA compared with the placebo group during the 6 months of supplementation. Messenger RNA (mRNA) of the Nuclear Factor kappa beta (NF-κB) and its target genes InterLeukin 1 beta (IL-1β) and IL-6 was downregulated significantly (p<0.05) in leukocytes from DMD boys supplemented with ω-3 long chain-PUFA for 6 months, compared to the placebo group. Omega-3 long chain-PUFA intake decreased the serum IL-1β (-59.5%; p=0.011) and IL-6 (-54.8%; p=0.041), and increased the serum IL-10 (99.9%, p<0.005), in relation to those with placebo treatment. Supplementation with ω-3 long chain-PUFA 2.9g/day is well-tolerated, has a beneficial reductive effect on proinflammatory markers, and increases an anti-inflammatory marker, indicating that ω-3 long chain-PUFA could have a potential therapeutic impact on chronic inflammation in DMD. This research is registered at clinicaltrials.gov (NCT018264229).

Deletion of the P2X4 receptor is neuroprotective acutely, but induces a depressive phenotype during recovery from ischemic stroke

IntroductionAcute ischemic injury leads to severe neuronal loss. One of the key mechanisms responsible for this effect is inflammation, which is characterized by the activation of myeloid cells, including resident microglia and infiltrating monocytes/macrophages. P2X4 receptors (P2X4Rs) present on these immune cells modulate the inflammatory response. For example, excessive release of adenosine triphosphate during acute ischemic stroke triggers stimulation of P2X4Rs, leading to myeloid cell activation and proliferation and further exacerbating post-ischemic inflammation. In contrast, during recovery P2X4Rs activation on microglia leads to the release of brain-derived neurotrophic factor (BDNF), which alleviate depression, maintain synaptic plasticity and hasten post-stroke behavioral recovery. Therefore, we hypothesized that deletion of the P2X4R specifically from myeloid cells would have differential effects on acute versus chronic recovery following stroke. MethodsWe subjected global or myeloid-specific (MS) P2X4R knock-out (KO) mice and wild-type littermates of both sexes to right middle cerebral artery occlusion (60min). We performed histological, behavioral (sensorimotor and depressive), and biochemical (quantitative PCR and flow cytometry) analyses to determine the acute (three days after occlusion) and chronic (30days after occlusion) effects of receptor deletion. ResultsGlobal P2X4R deletion led to reduced infarct size in both sexes. In MS P2X4R KO mice, only females showed reduced infarct size, an effect that did not change with ovariectomy. MS P2X4R KO mice of both sexes showed swift recovery from sensorimotor deficits during acute recovery but exhibited a more pronounced post-stroke depressive behavior phenotype that was independent of infarct size. Quantitative PCR analysis of whole cell lysate as well as flow-sorted myeloid cells from the perilesional cortex showed increased cellular interleukin 1 beta (IL-1β), interleukin 6 (IL-6), and tumor necrosis factor alpha (TNF-α) mRNA levels but reduced plasma levels of these cytokines in MS P2X4R KO mice after stroke. The expression levels of BDNF and other depression-associated genes were reduced in MS P2X4R KO mice after stroke. ConclusionsP2X4R deletion protects against stroke acutely but predisposes to depression-like behavior chronically after stroke. Thus, a time-sensitive approach should be considered when targeting P2X4Rs after stroke.