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Abstract Introduction: Pregnancy rhinitis is a common sex hormone-related otorhinolaryngological disorder. There are some epidemiological and physiological studies on pregnancy rhinitis, but histopathological and biomolecular changes have not been studied thoroughly. Objectives: The receptors VPAC1 and VPAC2 are known for their roles in allergic rhinitis. On the other hand, activation of subclinical allergy has been suggested in the pathophysiology of pregnancy rhinitis. Therefore, we aimed to compare the physiological and gestational pattern of VPAC1 and VPAC2 expression in rat nasal mucosa. Methods: Twenty adult Wister albino female rats were enrolled into the study. Two groups constituted as 10 control (group A) and 10 pregnant (group B) rats. They were fed ad libitum and sheltered at room temperature (22°±2°C). The rats were sacrificed at the 20th day of gestation by intraperitoneal injection of 400mg/kg Na-pentobarbitone. Then, 10 - 15 mL of blood was taken, and samples were reserved for the detection of serum estradiol and progesterone levels by ELISA test. The nasal septum was resected and divided in half for immunohistochemical analyses and real time polymerase chain reaction testing of VPAC1 and VPAC2. Results: VPAC1 and VPAC2 were found to be in all layers of septal specimens, but the immunostaining of surface epithelium was more distinct in specimens of both groups. We demonstrated higher overall staining intensity in the pregnant group. PCR revealed significant increase in expression of VPAC1 (p = 0.023) and VPAC2 (p = 0.021) in pregnant group when compared with control group. In addition, we demonstrated upregulatory effect of estradiol and progesterone on the vasoactive intestinal peptide receptor expression. Conclusions: Gestational up-regulation of nasal VPAC1 and VPAC2 was shown both by PCR and immunohistochemical analysis. These findings support the hypothesis that PR is caused by the activation of subclinical allergy that is present before pregnancy.
Resumo Introdução: A rinite gestacional é um distúrbio comum da otorrinolaringologia relacionado a hormônios sexuais. Existem alguns estudos epidemiológicos e fisiológicos sobre rinite gestacional, mas as alterações histopatológicas e biomoleculares ainda não foram estudadas completamente. Objetivo: Os receptores VPAC1 e VPAC2 são conhecidos por seu papel na rinite alérgica. Por outro lado, a ativação da alergia subclínica tem sido sugerida na fisiopatologia da rinite gestacional. Portanto, objetivamos comparar o padrão fisiológico e gestacional da expressão de VPAC1 e VPAC2 na mucosa nasal de ratos. Método: Vinte ratas fêmeas Wistar albinas adultas foram incluídas no estudo. Os dois grupos foram divididos em 10 ratas; controle (grupo A) e 10 ratas prenhes (grupo B). Elas foram alimentadas ad libitum e abrigadas em temperatura ambiente (22° ±2° C). Sacrificamos as ratas no 20° dia de gestação por injeção intraperitoneal de 400 mg/kg de sódio-pentobarbital. Em seguida, foram coletados 10 a 15 mL de sangue e as amostras foram reservadas para a detecção dos níveis séricos de estradiol e progesterona pelo método Elisa. O septo nasal foi ressecado e dividido em 2 para análises imuno-histoquímicas e testes de reação em cadeia da polimerase em tempo real, RT-PCR, de VPAC1 e VPAC2. Resultados: VPAC1 e VPAC2 foram encontrados em todas as camadas da amostra septal, mas a imunocoloração do epitélio de superfície foi mais distinta nas amostras de ambos os grupos. Demonstramos maior intensidade geral de coloração no grupo gestante. A reação de polimerase em cadeia revelou aumento significante na expressão de VPAC1 (p = 0,023) e VPAC2 (p = 0,021) no grupo gestante quando comparado ao grupo controle. Além disso, demonstramos um efeito up-regulador do estradiol e progesterona na expressão do receptor peptídeo intestinal vasoativo. Conclusão: A up-regulação gestacional dos receptores VPAC1 e VPAC2 nasais foi demonstrada tanto por reação de polimerase em cadeia quanto por análise imuno-histoquímica. Esses achados corroboram a hipótese de que a rinite gestacional é causada pela ativação de alergia subclínica presente antes da gestação.
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OBJECTIVES: TWIK-related potassium channel-1 (TREK-1) and Aquaporin 5 (AQP5) are involved in epithelial integrity and fluid transport, respectively. In this study, we aimed to compare physiological and gestational patterns of TREK-1 and AQP5 location and expression in rat larynx. Our secondary objective was to reveal the effect of estradiol (E2) and progesterone (PG) on these two biomolecules. METHODS: This study was conducted on 20 Wister albino female rats which were assigned as control (group A) and pregnant group (group B). The rats were sacrificed at 20th day of pregnancy. Blood was obtained directly from the ventricle for detection of serum E2 and PG levels. Larynx was resected for immunohistochemical analyses and real-time polymerase chain reaction testing for detection of TREK-1 and AQP5 staining and expression, respectively. RESULTS: Relative TREK-1 (P = 0.035) and AQP5 (P = 0.019) expression was found to be significantly high in group B when compared with group A. We found positive correlation between serum E2 levels and both biomolecules (TREK-1; P = 0.018, AQP5; P = 0.016). We also found positive correlation between serum PG levels and both biomolecules (TREK-1; P = 0.001, AQP5; P = 0.019). TREK-1 immunostaining was found to be higher in surface epithelium and lamina propria of vocal cord mucosa. AQP5 was particularly found to be located in basement membrane and adjacent superficial lamina propria. We revealed the physiological and gestational pattern of laryngeal TREK-1 and AQP5 expression for the first time. Gestational expression of both TREK-1 and AQP5 was found to be increased. Stimulatory effect of E2 and PG on laryngeal TREK-1 and AQP5 expression was also revealed. CONCLUSIONS: We revealed upregulatory effect of E2 and PG on laryngeal TREK-1 and AQP5 expression. Based on this finding, it can be suggested that TREK-1 and AQP5 play role in biomolecular processes leading gonadocorticoid-related voice changes.
Subject(s)
Aquaporin 5 , Potassium Channels, Tandem Pore Domain , Voice Disorders , Animals , Aquaporin 5/genetics , Aquaporin 5/metabolism , Female , Humans , Potassium Channels, Tandem Pore Domain/genetics , Pregnancy , Rats , Rats, WistarABSTRACT
INTRODUCTION: Pregnancy rhinitis is a common sex hormone-related otorhinolaryngological disorder. There are some epidemiological and physiological studies on pregnancy rhinitis, but histopathological and biomolecular changes have not been studied thoroughly. OBJECTIVES: The receptors VPAC1 and VPAC2 are known for their roles in allergic rhinitis. On the other hand, activation of subclinical allergy has been suggested in the pathophysiology of pregnancy rhinitis. Therefore, we aimed to compare the physiological and gestational pattern of VPAC1 and VPAC2 expression in rat nasal mucosa. METHODS: Twenty adult Wister albino female rats were enrolled into the study. Two groups constituted as 10 control (group A) and 10 pregnant (group B) rats. They were fed ad libitum and sheltered at room temperature (22°±2°C). The rats were sacrificed at the 20th day of gestation by intraperitoneal injection of 400mg/kg Na-pentobarbitone. Then, 10-15mL of blood was taken, and samples were reserved for the detection of serum estradiol and progesterone levels by ELISA test. The nasal septum was resected and divided in half for immunohistochemical analyses and real time polymerase chain reaction testing of VPAC1 and VPAC2. RESULTS: VPAC1 and VPAC2 were found to be in all layers of septal specimens, but the immunostaining of surface epithelium was more distinct in specimens of both groups. We demonstrated higher overall staining intensity in the pregnant group. PCR revealed significant increase in expression of VPAC1 (p=0.023) and VPAC2 (p=0.021) in pregnant group when compared with control group. In addition, we demonstrated upregulatory effect of estradiol and progesterone on the vasoactive intestinal peptide receptor expression. CONCLUSIONS: Gestational up-regulation of nasal VPAC1 and VPAC2 was shown both by PCR and immunohistochemical analysis. These findings support the hypothesis that PR is caused by the activation of subclinical allergy that is present before pregnancy.
Subject(s)
Hypersensitivity , Rhinitis , Animals , Estradiol , Female , Pregnancy , Progesterone , Rats , Rats, WistarABSTRACT
OBJECTIVE: The aim of this study is to reveal physiological expression and distribution of nuclear factor-kappa B (NF-κB) and MUC 5 subtype AC (MUC5AC) in rat laryngeal mucosa and to find out the effect of pregnancy and glucocorticoid treatment on these biomolecules. METHODS: This animal experiment was done in Experimental Animals Research and Application Center of Manisa Celal Bayar University in accordance with the accepted policy on the use of animals. A total of 30 young, adult Wister albino female rats were randomized into a control group (group A), a pregnant group (group B), and a steroid administered group (group C). Sacrification was done by injection of sodium-pentobarbitone (400 mg/kg) solution via intraperitoneal route in all groups. Serum estradiole (E2) and progesterone (PG) were determined by enzyme-linked immunosorbent assay. The relative expression and distribution of NF-κB and MUC5AC in laryngeal mucosa was studied both by immunohistochemistry (IHC) and polymerase chain reaction testing. Expression and immunohistochemical localization of NF-κB and MUC5AC was evaluated by light microscopy (Olympus BX41). In statistical analyses; relative expression of NF-κB and MUC5AC were compared on group basis. The effect of E2 and PG levels on these biomolecules was also evaluated. RESULTS: NF-κB was found to be significantly low both in group B (P < 0.05) and C (P < 0.001) when compared with group A, while MUC5AC was found to be significantly high both in group B (P < 0.05) and group C (P < 0.05) when compared with group A. Concerning IHC; NF-κB was found to be expressed in epithelium and lamina propria. MUC5AC was found to be expressed particularly in the epithelial layer in all groups. Statistically significant negative correlation between PG and NF-κB expression (P = 0.048), but no correlation between PG and MUC5AC expression (P = 0.487) were revealed. On the other hand, no correlation was found between E2 and the expression of relevant biomolecules (NF-κB [P = 0.270], MUC5AC [P = 0.829]). We also did found a significant negative correlation between the expression of NF-κB and MUC5AC (P = 0.031). CONCLUSIONS: In this study, the physiological expression of NF-κB and MUC5AC in rat laryngeal mucosa was shown for the first time both by polymerase chain reaction and IHC. The impact of pregnancy and glucocorticoid treatment on the expression and distribution of these biomolecules was also revealed. The expression of NF-κB was found to be decreased while the expression of MUC5AC was found to be increased both by pregnancy and glucocorticoid treatment. The inhibitory effect of serum PG on NF-κB expression in rat laryngeal mucosa was also shown for the first time. The expression of MUC5AC was found to be increased both in pregnant and glucocorticoid administered group. Negative correlation between NF-κB and MUC5AC expression was also revealed in rat larynx for the first time. These findings may partially unclose the histochemical background of voice changes caused by pregnancy and as well as by glucocorticoid treatment.
Subject(s)
Larynx , Mucin 5AC , Mucous Membrane , NF-kappa B , Animals , Female , Glucocorticoids/pharmacology , Glucocorticoids/therapeutic use , Mucin 5AC/genetics , Mucin 5AC/metabolism , NF-kappa B/metabolism , Pregnancy , RatsABSTRACT
OBJECTIVES: Mesenchymal stem cells are self-renewing stem cells. The human foreskin has potential to be used as a source of stem cells. The aim of the study was to obtain spheroid formation of human foreskin cells (hnFSSCs) isolated from newborn human foreskin tissue. In addition, the apoptotic and proliferative effects of a traditional plant, Corchorus olitorius L. (C. olitorius), on hnFSSC spheroids were investigated. MATERIALS AND METHODS: After a routine circumcision procedure the cells were isolated and cultured in suitable medium. The plant leaves was extracted with ethanol and their composition was analyzed by liquid chromatography coupled with mass spectrometry (LC-MS/MS). The foreskin stem cells were characterized immunocytochemically by CD45, CD34, and CD90 antibodies. hnFSSC spheroids were formed using the hanging drop technique. Immunofluorescence staining was used on the obtained spheroids to determine the distribution of caspase-3 and Ki-67 after being treated with C. olitorius extract for 48 h. RESULTS: Immunostaining analysis showed that hnFSSCs were positive for CD45 and CD34 and negative for CD90. According to LC-MS/MS C. olitorius was rich in flavanols and hydrocinnamic acid derivatives. Although the spheroids obtained were loose and floating, the cells interacted with each other. Caspase-3 activity was higher in the control group than in the extract-treated group and Ki-67 was higher in the extract-treated group than in the control group, suggesting that the plant might have the capacity to increase stem cell proliferation due to its rich polyphenolic content. CONCLUSION: The results suggest that hnFSSCs and spheroids might be used in stem cell generation, tissue repair and renewal as human foreskin tissue has potential to be used as a stem cell source. C. olitorius also increased proliferation of hnFSSCs, showing that polyphenols might increase proliferation of stem cells.
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Three-dimensional (3D) cancer tumor models are becoming vital approaches for high-throughput drug screening, drug targeting, development of novel theranostic systems, and personalized medicine. Yet, it is becoming more evident that the tumor progression and metastasis is fueled by a subpopulation of stem-like cells within the tumor that are also called cancer stem cells (CSCs). This study aimed to develop a tumoroid model using CSCs. For this purpose CD133+ cells were isolated from SaOS-2 osteosarcoma cell line with magnetic-activated cell sorting. To evaluate tumoroid formation ability, the cells were incubated in different cell numbers in agar gels produced by 3D Petri Dish® method. Subsequently, CD133+ cells and CD133- cells were co-cultured to investigate CD133+ cell localization in tumoroids. The characterization of tumoroids was performed using Live&Dead staining, immunohistochemistry, and quantitative polymerase chain reaction analysis. The results showed that, CD133+ , CD133- and SaOS-2 cells were all able to form 3D tumoroids regardless of the initial cell number, but, while 72 hr were needed for CD133+ cells to self-assemble, 24 hr were enough for CD133- and SaOS-2 cells. CD133+ cells were located within tumoroids randomly with high cell viability. Finally, when compared to two-dimensional (2D) cultures, there were 5.88, 4.14, 6.95, and 1.68-fold higher messenger RNA expressions for Sox2, OCT3/4, Nanog, and Nestin, respectively, in CD133+ cells that were cultured within 3D tumoroids, showing longer maintenance of stem cell phenotype in 3D, that can allow more relevant screening and targeting efficiency in pharmaceutical testing. It was concluded that CSC-based tumoroids are propitious as 3D tumor models to fill the gap between conventional 2D in vitro culture and in vivo animal experiments for cancer research.
Subject(s)
Models, Biological , Neoplastic Stem Cells , Osteosarcoma/metabolism , Spheroids, Cellular , AC133 Antigen/metabolism , Cell Line, Tumor , Humans , Neoplastic Stem Cells/chemistry , Neoplastic Stem Cells/cytology , Neoplastic Stem Cells/metabolism , Spheroids, Cellular/chemistry , Spheroids, Cellular/cytology , Spheroids, Cellular/metabolism , Tumor Cells, CulturedABSTRACT
BACKGROUND & OBJECTIVES: Vaginal atrophy is characterized by thinning of vaginal epithelial layers and decreased local blood flow. We aimed to evaluate the regenerative effects of Adipose derived mesenchymal stem cells (ADMSC) and Bone marrow derived mesenchymal stem cells (BMDSC) on vaginal atrophy in rat menopause model. MATERIALS AND METHODS: Rats were randomly divided into 4 (four) groups: sham, control, ADMSC, BMDSC. Vaginal epithelial thickness, structure of the lamina propria, blood vessels in the lamina propria, collagen deposition, and muscle structure were evaluated. Anti ER α, VEGF, VEGFR 1, Bax and bcl-2 antibodies were analyzed. Beta actin gene was used as endogenous control. Genetical differences among the groups were compared by using Kruskal Wallis and Mann Whitney U test. pâ¯<â¯0.05 was regarded as statistically significant. RESULTS: Epithelial thickness of ADMSC group was higher than control group, but less than sham group Epithelial thickness of BMDSC group was less than sham group. Lamina propria and muscle tissue of ADMSC and BMDSC groups were found to be similar to sham group. VEGFR-1, VEGF, Bax and ER-α staining levels were higher in ADMSC and BMDSC groups than control group. ADMSC group stained stronger with VEGFR-1 and VEGF than BMDSC group. Bcl-2 staining level was increased in ADMSC applied group. No statistically significant difference was detected in Bax and Bcl-2 genes and Bax-/Bcl-2 ratio. CONCLUSIONS: Although genetic expression might have ended and could not be significantly demonstrated, histological and immunohistochemical results favor ADMSC application in vaginal atrophy rather than BMDSC.
Subject(s)
Adipose Tissue/cytology , Biomarkers/metabolism , Bone Marrow Cells/cytology , Menopause/physiology , Mesenchymal Stem Cell Transplantation/methods , Vagina/pathology , Adipose Tissue/metabolism , Animals , Atrophy , Bone Marrow Cells/metabolism , Cells, Cultured , Disease Models, Animal , Estrogen Receptor alpha/metabolism , Female , Humans , Menopause/metabolism , Mesenchymal Stem Cells/cytology , Mesenchymal Stem Cells/metabolism , Rats , Vagina/metabolism , Vascular Endothelial Growth Factor A/metabolism , Vascular Endothelial Growth Factor Receptor-1/metabolism , bcl-2-Associated X Protein/metabolismABSTRACT
Cartilage created by tissue engineering is a promising new development in facial reconstructive surgery. The purpose of this study was to evaluate the histological results of implantation of synthetic polymer scaffold with chondrocytes differentiated from adipose-derived mesenchymal stem cells. Adipose tissue obtained from Wistar albino rats was dissociated, incubated and placed in culture medium. After a sufficient level of stem cell proliferation, the differentiation phase was started. Cells were collected on the 7th and 21st day of culture for chondrogenic characterization. After the 21st day of the differentiation phase of chondrocytes, they were transferred onto poly(dl-lactide-epsilon-caprolactone) synthetic polymer and culture continued for 24âhours. The scaffold with chondrocytes was then implanted into a subcutaneous area of skin on the back of the neck of the rat. Six weeks after implantation, all rats were sacrificed and the implantation areas were analyzed. Chondrocytes derived from adipogenic mesenchymal stem cells were stained positively with collagen II, aggrecan and Sox-9 after the differentiation stages. Histological examination of the excised material showed that chondrocytes were present, and the scaffold had been completely absorbed. The results of this study indicate that the differentiation method from mesenchymal stem cells to chondrogenic lineage was straightforward and scaffold with cells was easily accessible. This technique may be a good option for cartilage tissue engineering.
Subject(s)
Cartilage/growth & development , Chondrocytes/physiology , Chondrogenesis , Mesenchymal Stem Cells/physiology , Tissue Engineering/methods , Tissue Scaffolds , Adipose Tissue/cytology , Aggrecans/metabolism , Animals , Cartilage/cytology , Cartilage/metabolism , Cell Differentiation , Chondrocytes/cytology , Collagen Type II/metabolism , Male , Polymers , Rats , Rats, Wistar , SOX9 Transcription Factor/metabolismABSTRACT
OBJECTIVE: Colchicum pusillum belongs to the family Colchicaceae that particularly rich in tropolonic alkaloids. The aim of this study was to investigate the cytotoxicity and in vitro anticancer activity of Colchicum pusillum ethanolic extract on Colo-320 primer and Colo-741 metastatic colon adenocarcinoma cell lines. MATERIALS AND METHODS: Colchicum pusillum was collected and extracted with ethanol. Different concentrations of Colchicum pusillum extract were incubated for 24â¯h and 48â¯h with Colo-320 and Colo-741 cells. Cell growth and cytotoxicity were measured by 3(4,5dimethylthiazol2yl)2,5diphenyltetrazolium bromide (MTT) assays. Anticancer and antiproliferative activities of Colchicum pusillum were investigated by immunocytochemistry using antibodies directed against to ß-catenin, Ki-67, LGR-5 Ki-67, DKK1, Frizzled-4, Wnt4, Wnt7a and caspase3 in Colo-741 cells. RESULTS: All concentrations of Colchicum pusillum extract had toxic effect in Colo-320 cells. Because of this, we used Colchicum pusillum extract at 20⯵g/ml for evaluate anticancer activities only in Colo-741 cells. As a result of immunohistochemical staining, ß-catenin, LGR-5 and caspase-3 immunoreactivities were significantly increased while Wnt7a immunostaining intensity was decreased in Colo-741 cells. Conclusion We conclude that Colchicum pusillum extract increased ß-catenin and LGR-5 via Wnt/ß-catenin pathway in colon cancer cells. Interestingly, it decreased other signaling molecule, Wnt7a which is assumed to play protective role during carcinogenesis. Also, it increased significantly caspase-3 immunoreactivity showing that apoptotic pathways were triggered.
Subject(s)
Adenocarcinoma/metabolism , Antineoplastic Agents, Phytogenic/pharmacology , Colchicum/chemistry , Colonic Neoplasms/metabolism , Plant Extracts/pharmacology , Wnt Signaling Pathway/drug effects , Adenocarcinoma/drug therapy , Adenocarcinoma/pathology , Antineoplastic Agents, Phytogenic/chemistry , Cell Line, Tumor , Colonic Neoplasms/drug therapy , Colonic Neoplasms/pathology , Humans , Neoplasm Proteins/metabolism , Plant Extracts/chemistry , beta Catenin/metabolismABSTRACT
Introduction: Diabetic burn wounds and ulcers are significant complications of diabetic patients. The aim of this study is to investigate the use of platelet rich-plasma (PRP) and/or keratinocyte-like cells (KLCs) in diabetic thermal wound rat model and to evaluate EGF, FGF-2, TGF-ß1, COL1α2, MCP-1 and VEGF-α as wound healing markers at gene expression level. Method: In this study, we used adipose tissue as the source of mesenchymal stem cells (MSCs) and differentiated MSCs into KLCs. KLCs were characterized and transferred to the burn areas on the dorsum of streptozotocine (STZ)-induced diabetic rats. We prepared PRP from rat blood and evaluated its effect alone or in combination with KLCs. On 3rd, 7th, 10th and 14th days after treatment, wound areas were measured and biopsy samples were excised from the wound areas of the KLCs and/or PRP-treated and untreated diabetic rats to analyze gene expression levels of wound healing markers by qPCR. Results: We observed that, wound contraction started earlier in the PRP and/or KLCs-treated groups in comparison to the control group. However, PRP and KLCs when applied in combination showed additive affect in wound healing. In all groups treated with KLCs and/or PRP, the gene expression levels of evaluated growth factors and COL1α2 increased, while MCP-1 levels decreased when compared to the untreated diabetic rats. In addition, the most prominent difference in qPCR results belongs to combined PRP and KLCs-treated group. Conclusion: We demonstrated that applying PRP and KLCs in combination has a greater potential for treatment of diabetic burn wounds.
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Research using animal models gives human trials hope for recovery in many fields of regenerative medicine, although they are sometimes poor predictors for human experiences. Our goal was to investigate whether rat chondrocytes, differentiated from adipose-derived stem cells, could be transplanted using a new, easily shaped, bioactive glass scaffold, and to show the immunohistochemical results. Intraperitoneal and retroperitoneal adipose tissue was extracted from 6 male Wistar albino type rats. The fatty tissue samples were fragmented and incubated. Chondrogenic differentiation was carried out and collagen type II, bFGF, and Sox-9 immunohistochemical characterization analysis was performed. Differentiated chondrocytes were implanted on 13-93B3 bioactive glass scaffolds and transplanted into the right ears of the rats. As control, only the biomaterial was transplanted into the left ears of the rats. After 1 month, the rats were sacrificed and transplantation areas were examined immunohistochemically. Histological examination of control samples from the left ears revealed that the biomaterial was covered with connective tissue, its general structure was preserved, and resorption of the scaffold had started. In specimens from the right ears, the biomaterial was covered with connective tissue, its structure was preserved, cartilage cells were present around the biomaterial, and the presence of cartilage tissue was demonstrated immunohistochemically. In conclusion, 13-93B3 bioactive glass scaffold contributed to the formation of new collagen and the survival of chondrocytes, and is a promising new biomaterial that will prove very useful in regenerative medicine.
Subject(s)
Biocompatible Materials , Cartilage/metabolism , Chondrocytes/transplantation , Chondrogenesis , Tissue Scaffolds , Adipose Tissue/cytology , Animals , Cartilage/growth & development , Cell Differentiation , Collagen Type II/metabolism , Fibroblast Growth Factor 2/metabolism , Male , Rats , Rats, Wistar , SOX9 Transcription Factor/metabolism , Stem Cells , Tissue Scaffolds/chemistryABSTRACT
AIM: To contribute to the understanding of the pathogenesis of chronic spontaneous urticaria (CSU) by identifying its relationship with autoimmunity and cytokines using the autologous serum skin test (ASST) and peripheral blood mononuclear cell culture (PBMC) method. MATERIAL AND METHODS: Interleukins (IL)-4, IL-10, transforming growth factor (TGF-ß1), interferon (IFN)-γ, IL-17A, and IL-23 levels in cell supernatants obtained by the PBMC method were measured using ELISA. Disease activity was assessed by determining the urticaria activity score (UAS). RESULTS: A total of 40 patients diagnosed with CSU participated in this study. Twenty patients had positive ASST results, and 20 had negative results. The control group included 20 healthy volunteers. We found that the IL-23 (p = 0.01), IL-10 (p = 0.04) and IL-4 (p = 0.04) levels of the patient groups were significantly lower compared with those of the control group. The IL-23 (p = 0.009), IL-10 (p = 0.009), IL-4 (p = 0.001), and IL-17 (p = 0.05) levels of the ASST(-) patient group were significantly lower compared with those of the control group. In addition, the IL-4 (p = 0.03) and IFN-γ (p = 0.05) levels of the ASST(+) patient group were significantly lower compared with those of the control group, and the ASST(+) patients had a significantly higher UAS than the ASST(-) patients (p = 0.021). CONCLUSIONS: These results, when considered together with current reports in the literature, indicate that immune dysregulation occurs in the pathogenesis of CSU, causing cytokine imbalance.
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CONTEXT: Almond oil is used in traditional and complementary therapies for its numerous health benefits due to high unsaturated fatty acids content. OBJECTIVES: This study investigated the composition and in vitro anticancer activity of almond oil from Northern Cyprus and compared with almond oil from Turkey. MATERIALS AND METHODS: Almond oil from Northern Cyprus was obtained by supercritical CO2 extraction and analyzed by GC-MS. Almond oil of Turkey was provided from Turkish pharmacies. Different concentrations of almond oils were incubated for 24 and 48 h with Colo-320 and Colo-741 cells. Cell growth and cytotoxicity were measured by MTT assays. Anticancer and antiprolifetarive activities of almond oils were investigated by immunocytochemistry using antibodies directed against to BMP-2, ß-catenin, Ki-67, LGR-5 and Jagged 1. RESULTS: Oleic acid (77.8%; 75.3%), linoleic acid (13.5%; 15.8%), palmitic acid (7.4%; 6.3%), were determined as the major compounds of almond oil from Northern Cyprus and Turkey, respectively. In the MTT assay, both almond oils were found to be active against Colo-320 and Colo-741 cells with 1:1 dilution for both 24 h and 48 h. As a result of immunohistochemical staining, while both almond oils exhibited significant antiproliferative and anticancer activity, these activities were more similar in Colo-320 cells which were treated with Northern Cyprus almond oil. DISCUSSION AND CONCLUSION: Almond oil from Northern Cyprus and Turkey may have anticancer and antiproliferative effects on colon cancer cells through molecular signalling pathways and, thus, they could be potential novel therapeutic agents.
Subject(s)
Antineoplastic Agents, Phytogenic/pharmacology , Colonic Neoplasms/pathology , Fatty Acids/pharmacology , Plant Oils/pharmacology , Prunus dulcis , Seeds , Antineoplastic Agents, Phytogenic/isolation & purification , Antineoplastic Agents, Phytogenic/therapeutic use , Cell Line, Tumor , Cell Survival/drug effects , Cell Survival/physiology , Colonic Neoplasms/drug therapy , Dose-Response Relationship, Drug , Fatty Acids/isolation & purification , Fatty Acids/therapeutic use , Humans , Plant Oils/isolation & purification , Plant Oils/therapeutic useABSTRACT
Objective: To compare the distribution of proopiomelanocortin (POMC) in decidua and placenta samples from missed abortion and voluntary termination cases in order to research the effects in the etiology of missed abortion. Study Design: Decidual materials were collected from patients who were diagnosed with missed abortion (n=19) and legal voluntary termination cases (n=15) under 10 gestational weeks. Materials were divided into 2 groups for examination. For all samples, POMC primary antibody was performed by immunohistochemical staining. The number of stained cells was calculated by using the H-score technique. Results: In the missed abortion group the mean age was 28.7 (1841), and in the control group the mean age was 27.5 (2137). POMC immunoreactivity was determined to be lower in the parenchyma and placenta of the missed abortion group than those of the control group. POMC immunoreactivities were found to be higher in both the syncytiotrophoblast and cytotrophoblast cells of the missed abortion group than those of the control group (p<0.005). Conclusion: POMC has become a paradigmatic polypeptide precursor and has a role in the parturition process. Local production of POMC in placenta and decidua may influence pregnancy and may have a role in missed abortion pathogenesis.
Subject(s)
Abortion, Missed/metabolism , Abortion, Spontaneous/metabolism , Decidua/metabolism , Pro-Opiomelanocortin/metabolism , Abortion, Spontaneous/prevention & control , Adult , Case-Control Studies , Female , Humans , Immunohistochemistry , Neovascularization, Pathologic/metabolism , Placenta/pathology , Pregnancy , Pregnancy Trimester, First/metabolism , Trophoblasts/pathology , Young AdultABSTRACT
Hypoxia is a pathalogical condition in which tissues are deprived of adequate oxygen supply. The hypoxia effect on tumors has a critically important role on maintenance of cancer stem cell phenotype. The aim of this study is to investigate the effects of hypoxia on cancer stem cells on three dimensional (3D) in vitro culture models. Osteosarcoma stem cells characterized by CD133 surface protein were isolated from osteosarcoma cell line (SaOS-2) by magnetic-activated cell sorting (MACS) technique. Isolated CD133(+) and CD133(-) cells were cultivated under hypoxic (1% O2) and normoxic conditions (21% O2) for 3 days. For the 3D model, bacterial cellulose scaffold was used as the culture substrate. 3D morphologies of cells were examined by scanning electron microscopy (SEM); RT-PCR and immunocytochemistry staining were used to demonstrate conservation of the cancer stem cell phenotype in 3D environment under hypoxic conditions. Cell viability was shown by MTT assay on 3. and 7. culture days. This study is seen as an introduction to develop a 3D hypoxic cancer stem cell based tumor model to study CSC behavior and tumor genesis in vitro.
Subject(s)
Neoplastic Stem Cells/pathology , Osteosarcoma/pathology , Stem Cell Niche , AC133 Antigen/metabolism , Cell Count , Cell Hypoxia , Cell Line, Tumor , Cell Shape , Humans , Immunohistochemistry , Immunomagnetic Separation , Phenotype , RNA, Messenger/genetics , RNA, Messenger/metabolism , Real-Time Polymerase Chain ReactionABSTRACT
OBJECTIVES: To determine the efficacy of montelukast in comparison with cabergoline in the prevention of ovarian hyperstimulation syndrome (OHSS) in rats. MATERIAL AND METHODS: An experimental OHSS model was formed in 35 female Wistar rats. Rats (22 days old) were randomized into 5 groups, each containing 7 animals. The control group received no therapy; the mild OHSS group was administered pregnant mare serum gonadotropin (PMSG) 10 IU for 4 days, hCG 10 IU on the 5th day; the severe OHSS group received PMSG 10 IU for 4 days, hCG 30 IU on the 5th day The montelukast group: received montelukast 10 mg/kg/day and the cabergoline group was administered cabergollne 100 microg/kg/day via oral gavage for 6 days (days 22-27), in addition to those of severe OHSS. All groups were sacrificed on 28th day Body weight, ovarian diameter and weight, vascular permeability vascular endothelial growth factor (VEGF), semiquantitative VEGF receptor-1, and VEGF receptor-2 (VEGFR-2) immunohistochemistry were evaluated. RESULTS: Ovarian diameter and VEGF expression were significantly lower in the montelukast and cabergoline groups than in the severe OHSS group. While montelukast was more effective in limiting vascular permeability in the severe OHSS, cabergoline was superior to montelukast with respect to the limiting effect on increased body weight and VEGFR-2 expression. CONCLUSIONS: The VEGF/VEGFR-2 interaction plays an important role in OHSS pathogenesis. Montelukast limits VEGF expression, and cabergoline reduces both VEGF and VEGFR-2 expressions; they are both effective therapies for the prevention of severe OHSS.
Subject(s)
Acetates/pharmacology , Disease Models, Animal , Ergolines/pharmacology , Ovarian Hyperstimulation Syndrome/drug therapy , Quinolines/pharmacology , Acetates/administration & dosage , Animals , Cabergoline , Chorionic Gonadotropin/pharmacology , Cyclopropanes , Dose-Response Relationship, Drug , Ergolines/administration & dosage , Female , Ovarian Hyperstimulation Syndrome/prevention & control , Ovary/drug effects , Quinolines/administration & dosage , Random Allocation , Rats , Rats, Wistar , SulfidesABSTRACT
OBJECTIVE: To identify the role of furin, TNF-α, and TGF-ß2 in human missed abortion pathogenesis. STUDY DESIGN: Decidual materials were collected from patients diagnosed with a missed abortion (n = 10) (missed abortion group) and from legal voluntary termination cases at < 10 gestational weeks (n = 10) (normal pregnancy group). Tissue samples were collected from each group by dilation and curettage under mask anesthesia. For all tissue samples,furin, TNF-α, and TGF-ß2 primary antibodies were performed by immunohistochemical staining. The number of stained cells was evaluated by using the H-score technique. RESULTS: In immunohistochemical examination, the immunoreactivities of furin, TNF-α, and TGF-ß2 were found to be higher in syncytiotrophoblastic cells in the missed abortion group than in the normal pregnancy group (p < 0.005). Additionally, high immunoreactivity of TNF-α and TGF-ß2 molecules was established only in cytotrophoblastic cells of missed abortions (p < 0.005) in examination at decidual cells of the missed abortion group; furin immunoreactivities were detected higher in the missed abortion group than in the control group, but TNF-α and TGF-ß2 immunoreactivity were increased in number in the normal pregnancy group (p < 0.005). CONCLUSION: It is considered that high levels offurin and the 2 furin-related proteins (TNF-α and TGF-ß2), which play important roles in proliferation, invasion, migration, differentiation, and survival of cells, may be the reason of proceeding decidualization, placentation, and prevention from abortion, in spite of terminating thefetal life.
Subject(s)
Abortion, Induced , Abortion, Missed/metabolism , Endometrium/metabolism , Furin/biosynthesis , Lymphotoxin-alpha/biosynthesis , Tumor Necrosis Factor-alpha/biosynthesis , Adult , Female , Furin/analysis , Humans , Immunohistochemistry , Lymphotoxin-alpha/analysis , Pregnancy , Pregnancy Trimester, First , Tumor Necrosis Factor-alpha/analysis , Young AdultABSTRACT
Male infertility is correlated with sperm morphology and sperm DNA damage, which are completely different from that of fertile individuals. An accurate sperm DNA damage analysis and ultrastructural examination of the ejaculate provide important support in the clinical evaluation. It is supposed that in the near future, the fertilization rate, pregnancy rate, and miscarriages could be predicted using the combination of these types of tests in assisted reproductive technologies (ARTs). For this purpose, we report a very rare case of an infertile man having short tail sperm. The infertile man and his wife underwent in vitro fertilization (IVF) with intracytoplasmic sperm injection (ICSI). During this process, we examined the ultrastructure of the ejaculated sperm with transmission electron microscopy (TEM) and calculated the sperm DNA damage with terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) and COMET assays. Then, we evaluated the association between sperm DNA damage and embryo quality.
ABSTRACT
OBJECTIVE: Heparin-binding epidermal growth factor (HB-EGF) has pleiotropic biological functions in the female reproductive tract. HB-EGF has a function in the menstruation cycle, implantation, decidualization, placenta development, and also inhibition of apoptosis. This study aims to investigate a possible role of HB-EGF in missed abortion. MATERIALS AND METHODS: Decidual and placental tissue samples were obtained from women with unwanted pregnancy as the control group and from women with missed abortions as the patient group. Immunohistochemistry was utilized to compare HB-EGF expression of fibroblast and decidual cells in uterine decidual stroma and fibroblasts and mesenchymal cells in placental villous stroma; the TUNEL technique was used to detect apoptotic cells within the decidual and placental tissues of the two groups. RESULTS: It was demonstrated that HB-EGF expression in both uterine decidual stroma and placenta stroma was increased in the missed abortion group (142.70 ± 12.80; 116.10 ± 14.16, respectively), compared with the normal pregnancy group (101.60 ± 14.18; 81.60 ± 10.74, respectively). It was also shown that there was no difference in TUNEL (Terminal deoxynucleotide transferase dUTP Nick End Labelling) positive cells between the uterine decidual stroma (11.4 ± 3%; 13.6 ± 3%, respectively), placental villous stroma (13.7 ± 3%; 15.9 ± 3%, respectively), and cytotrophoblast-syncytiotrophoblast cells (7.3 ± 2; 9.8 ± 3, respectively) of the two groups. CONCLUSION: This data supports the hypothesis that increased HB-EGF expression in a missed abortion may prevent the discharge of the dead fetus.
Subject(s)
Abortion, Missed/metabolism , Decidua/metabolism , Heparin-binding EGF-like Growth Factor/metabolism , Placenta/metabolism , Abortion, Missed/diagnosis , Adolescent , Adult , Apoptosis , Decidua/pathology , Female , Fibroblasts/metabolism , Humans , Immunohistochemistry , In Situ Nick-End Labeling , Placenta/pathology , Pregnancy , Young AdultABSTRACT
OBJECTIVE: To investigate the possible protective effect of a single dose of ketamine and the synergistic effect between ketamine and 2-mercaptoethane sulfonate (mesna) against ifosfamide-induced hemorrhagic cystitis. MATERIALS AND METHODS: 35 adult female wistar rats were divided into five groups and pretreated with ketamine at 10 mg/kg and/or mesna 400 mg/kg 30 minutes before intraperitoneal injection of IFS (400 mg/kg) or with saline (control group). Hemorrhagic cystitis was evaluated 24 hours after IFS injection according to bladder wet weight (BWW), and microscopic changes, i.e. edema, hemorrhage, cellular infiltration, and urothelial desquamation. The markers of oxidative damage including nitric oxide (NO) and malondialdehyde (MDA) levels and the expressions of tumor necrosis factor alpha (TNF-α), interleukin 1-beta (IL-1ß), inducible nitric oxide synthase (i-NOS) and endothelial nitric oxide synthase (e-NOS) were also assayed in the bladder tissues. RESULTS: Pretreatment with ketamine alone or ketamine in combination with mesna reduced the IFS-induced increase of BWW (58,47% and 63,33%, respectively, P < 0.05). IFS- induced microscopic alterations were also prevented by ketamine with or without mesna (P < 0.05). In addition, also statistically insignificant, the bladder tissue expressions of IL-1ß were lower in ketamine and/or mesna-receiving groups (P > 0,05). The parameters of oxidative stress, the NO and the MDA contents of the bladder tissues of the study groups were not different. CONCLUSION: The results of the present study suggest that a single dose of ketamine pretreatment attenuates experimental IFS-induced bladder damage. It is therefore necessary to investigate ketamine locally and systematically with various dosing schedules in order to reduce the bladder damage secondary to oxazaphosphorine-alkylating agents and these results may widen the spectrum of ketamine.