Platelet-rich plasma and platelet-derived lipid factors induce different and similar gene expression responses for selected genes related to wound healing in rat dermal wound environment.

Although platelet-rich plasma (PRP) is the plasma fraction that contains higher levels of platelet-sequestered proteins such as growth factors and chemokines, it is also abundant in bioactive lipids whose role in wound healing has not been well characterized. This study provides a preliminary evaluation for the effect of the lipid component of PRP on selected genes related to wound healing. Sprague-Dawley rats were classified into four groups after induction of full thickness excisional wounds: the lipid fraction (LF) (lipid extract from PRP) group, PRP group, dimethyl sulfoxide group, and sham group. Subsequently, relevant groups were topically treated with test preparations. Healing wounds were collected on 3rd, 7th and 14th days, and expression levels of 12 genes were determined using qPCR. LF treatment-induced gene expression signature distinct from that induced by PRP treatment, although there are some overlaps in LF- and PRP-responsive genes. Differentially expressed all eight genes (Cxcl5, Cxc11, Egfr, Tgfb1, IL10, Tgfa, Mmp1, and Mmp7) to LF response were significantly down-regulated at either 3rd, 7th, or 14th days. Also, the comparison between LF- and PRP-treatment groups showed that the LF significantly decreased expression of Cxcl11, Mmp7, and Tgfa mRNA on day 7 of healing. This study revealed that PRP and its LF induced different and similar gene expression responses of the skin during the repair of full thickness excisional wounds. Identifying mRNA response to LF treatment at whole transcriptome level can be beneficial for comprehensive understanding of the role of platelet-derived lipid factors in wound healing processes.

Evaluation of the in vivo wound healing potential of the lipid fraction from activated platelet-rich plasma.

Previous in vitro studies suggest a direct relevance for the peptide-free lipid fraction (LF) of platelet-rich plasma (PRP) in biological mechanisms related to wound healing. However, there are no scientific reports to date on the wound healing activities of this lipid component in vivo. Thus, the present study provides a scientific evaluation for the wound healing potential of the lipid portion of the activated PRP. For the wound healing activity assessment, in vivo full-thickness excisional wounds were created on the dorsal skin of Sprague-Dawley rats. Lipid extract from pooled PRP was applied topically to the wounds on 0, 3, and 7 days after injury. Histological assessment of epidermal and dermal regeneration, granulation tissue thickness and angiogenesis by Sirius red and Masson's trichrome staining, in addition to immunohistochemical staining for transforming growth factor beta-1 (TGF-ß1), collagen type I (COL I), and collagen type III (COL III) were performed on skin biopsies at 3, 7 and 14 days. The total histological scores of the LF group were significantly higher than the 25% dimethylsulfoxide-control group. According to the immunohistochemical staining, the observed expression changes for TGF-ß1, COL I and III at 3, 7, and 14 days after wounding were significantly better in the study group than the control group. Furthermore, COL I/III ratio in the lipid extract-treated group at day 14 was much higher than that of the control group. Meanwhile, analysis of the data also indicated that the LF has less positive effects on all evaluated parameters than PRP. From the present data, it could be concluded that the peptide-free LF of PRP has potent wound healing capacity in vivo for cutaneous wounds, although not as much as that of PRP. Strengthening our understanding of the wound healing potential of lipid components of PRP and platelet-derived lipid factors may provide new avenues for improving the healing process of a wound with elevated protease activity.

Anesthesia may alter mRNA expression of certain wound healing-associated genes in dermal wound environment of the rats.

Some anesthetics including ketamine/xylazine and thiopental have been shown to alter the expression of genes related with inflammatory cytokines and chemokines in previous studies unassociated with wound healing, arising the question of whether commonly used anesthetics in wound healing models could interfere with the transcriptional responses of the genes associated with skin wound healing. The gene expression profile in wound biopsies of rats who received widely used anesthetics doses of intraperitoneal ketamine/xylazine (50 mg/kg and 10 mg/kg) or thiopental (50 mg/kg) in comparison with control rats was analyzed by monitoring the expression of genes effective on various phases of wound healing. The expression levels of 84 genes were determined on 3rd, 7th and 14th days of post-wounding using a qPCR array system. Of the genes either up or downregulated fivefolds or more, three (Egf, Col5a1 and Cxcl3) and two (Tgfa and Il2) genes were found to be the most responsive ones to ketamine/xylazine or thiopental anesthesia respectively in a period of 14 days after correction for multiple testing. However, up to 22 and 24 genes for ketamine/xylazine and thiopental were found to be differentially expressed in the same period without correction for multiple-comparisons testing (p < 0.05). In conclusion, our data suggest that ketamine/xylazine and thiopental may alter the transcriptional responses of some genes associated with wound healing in rats. We strongly suggest to consider the possible alteration effect of these anesthetics on gene expression in animal models of dermal wound healing.

NFKB1 rs28362491 and pre-miRNA-146a rs2910164 SNPs on E-Cadherin expression in case of idiopathic oligospermia: A case-control study.

BACKGROUND: A notable proportion of idiopathic male infertility cases is accompanied by oligozoospermia; and yet, the molecular mechanisms of fertilization problem underlying this defect are still unclear. Epithelial cadherin has been involved in several calcium-dependent cell-to-cell adhesion events; however, its participation in gamete interaction has also not been fully investigated. OBJECTIVE: The aim was to investigate the changes in the expression of E-cadherin, based on the frequency of Single nucleotide polymorphisms in Nuclear Factor Kappa-B 1 and pre-mir-146a in oligospermic men. MATERIALS AND METHODS: In this case-control study, semen and blood samples of 131 oligospermic men as the case group and 239 fertile healthy men as the control group were analyzed. Variants single nucleotide polymorphisms rs28362491 and rs2910164 were performed using polymerase chain reaction-restriction fragment length polymorphism method and E-cadherin expression were determined by immunoprecipitation studies. RESULTS: ins/ins genotype of rs28362491 was determined as a risk factor for idiopathic oligospermia by 1.73 times (p=0.0218), whereas no significant differences were found between the groups concerning pre-mir-146a rs2910164 polymorphism (p=0.2274 in case of GC genotype and p=0.9052 in case of GG genotype). Combined genotype analysis results did not show any notable differences between the multiple comparisons of 28362491-rs2910164 in oligospermic men and control groups. In addition, E-cadherin expression of oligospermic men with ins/ins genotype was significantly lower than patients with del/ins genotype (p=0.0221). E-cadherin expression level was low in oligospermic men with respect to the control group in presence of ins/ins genotype of NFKB1 gene. CONCLUSION: These results suggest that ins allele prevents binding of surface proteins to spermatozoa, leading to a low affinity of sperm-oocyte interaction in oligospermic men.

Genetic Variation in NFKB1 Gene Influences Liver Enzyme Levels in Morbidly Obese Women.

BACKGROUND: Morbid obesity (MO), characterized by low-grade inflammation, is associated with increased C-reactive protein (CRP). NF-KB is a candidate factor for inflammatory responses in inflammatory diseases such as obesity. The objective of our study was to investigate the relationship between NFKB1 gene variations and the risk of MO in the context of the high/normal level of liver enzymes such as Alanine aminotransferase (ALT), Aspartate aminotransferase (AST), gamma-glutamyl transpeptidase (GGT), and alkaline phosphatase (ALP). METHODS: We analyzed the distribution of NFKB1 -94 ins/del ATTG (rs28362491) polymorphism using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP)and liver enzymes serum levels using ELISA in 182 MO patients with CRP level ≥20 mg/L and 200 healthy controls in a female Turkish population. RESULTS: We found that having ins/ins genotype of rs28362491 is a risk factor in both high level and normal level liver enzymes of ALT (P = 0.0335, P = 0.0134), AST (P = 0.0285, P = 0.0113) and ALP (P = 0.0079, P = 0.0363) whereas having ins/ins genotype of rs28362491 is a risk factor in only high-level liver enzyme of GGT (P = 0.0003). CONCLUSION: Our results suggest that ins/ins genotype of SNP rs28362491 is linked to MO with high-level ALT, AST, ALP, and GGT.

Gene expression profiling of Lucilia sericata larvae extraction/secretion-treated skin wounds.

The larvae of Lucilia sericata have been successfully used as medicinal maggots in the healing of wounds. The excretion/secretion (ES) products of the larvae have been shown to efficiently debride wounds and help the healing process. The mechanisms underlying ES-induced wound healing are not yet completely understood. One of the intriguing questions is the role of ESs in modulating gene expression at the transcriptional level in the skin wound environment during the healing process. To address this question, a study was conducted in which the ES-induced gene expression profile in wound biopsies and ES-treated wounds of rat skin in comparison with control group was analyzed at the molecular level by monitoring the expression of genes associated with wound healing. The expression levels of 82 genes at 4, 7, and 10 days after wounding were determined using a PCR array system following cDNA synthesis. A comparison from wounds revealed that 38 mRNAs (≥5-fold expression) were differentially expressed in the ES-treated skin. For 27 genes, the multiple-test corrected p-value was statistically significant (p≤0.00061). The expression pattern of these mRNAs was also altered during a period of 10 days. Many of the upregulated or downregulated mRNAs with therapy were extracellular matrix, cell adhesion-related proteins and growth factors. The genes that have the highest fold change (>1000-fold) were Col1a2, Col4a1, Ctsk, Ccl7, Angpt1, Cd40lg, Egf and Itgb5. Several of these gene products might play key roles in ES-induced wound healing. These findings may provide new insight for an understanding of the therapeutic potential of ESs for wound healing.