Displaying publications 1 - 20 of 98 in total

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  1. Giribabu N, Eswar Kumar K, Swapna Rekha S, Muniandy S, Salleh N
    PMID: 25852767 DOI: 10.1155/2015/542026
    The effect of V. vinifera seeds on carbohydrate metabolizing enzymes and other enzymes of the liver in diabetes is currently unknown. We therefore investigated changes in the activity levels of these enzymes following V. vinifera seed extract administration to diabetic rats. Methods. V. vinifera seed ethanolic extract (250 and 500 mg/kg/day) or glibenclamide (600 μg/kg/day) was administered to streptozotocin-induced male diabetic rats for 28 consecutive days. At the end of treatment, liver was harvested and activity levels of various liver enzymes were determined. Levels of thiobarbituric acid reactive substances (TBARS) were measured in liver homogenates and liver histopathological changes were observed. Results. V. vinifera seed ethanolic extract was able to prevent the decrease in ICDH, SDH, MDH, and G-6-PDH and the increase in LDH activity levels in liver homogenates. The seed extract also caused serum levels of ALT, AST, ALP, ACP, GGT, and total bilirubin to decrease while causing total proteins to increase. Additionally, the levels of ALT, AST, and TBARS in liver homogenates were decreased. Histopathological changes in the liver were reduced. Conclusion. Near normal activity levels of various enzymes and histology of the liver following V. vinifera seed ethanolic extract administration may be due to decrease in liver oxidative stress in diabetes.
  2. Kamarudin NH, Jalil AA, Triwahyono S, Artika V, Salleh NF, Karim AH, et al.
    J Colloid Interface Sci, 2014 May 1;421:6-13.
    PMID: 24594025 DOI: 10.1016/j.jcis.2014.01.034
    Mesoporous silica nanoparticles (MSNs) were synthesized with variable microwave power in the range of 100-450 W, and the resulting enhancement of MSN crystal growth was evaluated for the adsorption and release of ibuprofen. X-ray diffraction (XRD) revealed that the MSN prepared under the highest microwave power (MSN450) produced the most crystallized and prominent mesoporous structure. Enhancement of the crystal growth improved the hexagonal order and range of silica, which led to greater surface area, pore width and pore volume. MSN450 exhibited higher ibuprofen adsorption (98.3 mg/g), followed by MSN300(81.3 mg/g) and MSN100(74.1 mg/g), confirming that more crystallized MSN demonstrated higher adsorptivity toward ibuprofen. Significantly, MSN450 also contained more hydroxyl groups that provided more adsorption sites. In addition, MSN450 exhibited comparable ibuprofen adsorption with conventionally synthesized MSN, indicating the potential of microwave treatment in the synthesis of related porous materials. In vitro drug release was also investigated with simulated biological fluids and the kinetics was studied under different pH conditions. MSN450 showed the slowest release rate of ibuprofen, followed by MSN300 and MSN100. This was due to the wide pore diameter and longer range of silica order of the MSN450. Ibuprofen release from MSN450 at pH 5 and 7 was found to obey a zero-order kinetic model, while release at pH 2 followed the Kosmeyer-Peppas model.
  3. Hassan NS, Jalil AA, Fei ICM, Razak MTA, Khusnun NF, Bahari MB, et al.
    Chemosphere, 2023 Oct;338:139502.
    PMID: 37453521 DOI: 10.1016/j.chemosphere.2023.139502
    Vanadia (V2O5)-incorporated fibrous silica-titania (V/FST) catalysts, which were successfully synthesized using a hydrothermal method followed by the impregnation of V2O5. The catalysts were then characterized using numerous techniques, including X-ray diffraction, field emission scanning electron microscopy, transmission electron microscopy, nitrogen adsorption-desorption analyses, ultraviolet-visible diffuse reflectance spectroscopy, Fourier-transform infrared, X-ray photoelectron spectroscopy, and photoluminescence (PL) analyses. The study found that varying the amount of V2O5 (1-10 wt%) had a significant impact on the physicochemical properties of the FST, which in turn improved the photodegradation efficiency of two organic compounds, ciprofloxacin (CIP) and congo red (CR). 5V/FST demonstrated the best performance in degrading 10 mg L-1 of CIP (83%) and CR (100%) at pH 3 using 0.375 g L-1 catalyst under visible light irradiation within 180 min. The highest photoactivity of 5V/FST is mainly due to higher crystallinity and the highest number of V2O5-FST interactions. Furthermore, as demonstrated by PL analysis, the 5V/FST catalyst has the most significant impact on interfacial charge transfer and reduces electron-hole recombination. The photodegradation of both contaminants follows the Langmuir-Hinshelwood pseudo-first-order model, according to the kinetic study. The scavenger investigation demonstrated that hydroxyl radicals and holes dominated species in the system, indicating that the catalyst effectively generated reactive species for pollutant degradation. A possible mechanism was also identified for FST and 5V/FST. Interestingly, V2O5 acts as an electron-hole recombination inhibitor on FST for selective hole oxidation of ciprofloxacin and congo red photodegradation. Finally, the degradation efficiency of the catalyst remained relatively stable even after five cyclic experiments, indicating its potential for long-term use in environmental remediation.
  4. Karim K, Giribabu N, Muniandy S, Salleh N
    J. Membr. Biol., 2016 04;249(1-2):65-76.
    PMID: 26403527 DOI: 10.1007/s00232-015-9848-z
    We hypothesized that progesterone-induced decrease in uterine fluid pH involves V-ATPase. In this study, expression and functional activity of V-ATPase in uterus were investigated under progesterone influence. Ovariectomized adult female rats received subcutaneous injection of estradiol-17β (1 µg/kg/day) or progesterone (20 mg/kg/day) for 3 days or 3 days estradiol-17β followed by 3 days vehicle, progesterone, or estradiol-17β plus progesterone. Mifepristone, a progesterone receptor blocker, was concomitantly given to the rats which received progesterone. A day after last injection, rate of uterine fluid secretion, its HCO3 (-) concentration, and pH were determined via in vivo uterine perfusion in rats under anesthesia. V-ATPase inhibitor, bafilomycin, was introduced into the perfusion buffer, and changes in these parameters were observed. Expression of V-ATPase A1 and B1/2 proteins and mRNAs in uterus were quantified by Western blotting and real-time PCR, respectively. Distribution of these proteins was observed by immunohistochemistry. Our findings showed that under progesterone influence, uterine fluid secretion rate, HCO3 (-) concentration, and pH were significantly reduced. Administration of bafilomycin did not cause significant changes in fluid secretion rate; however, HCO3 (-) concentration and pH were significantly elevated. In parallel with these changes, expression of V-ATPase A1 and B1/2 proteins and mRNAs were significantly increased with these proteins highly distributed in uterine luminal and glandular epithelia. In conclusion, increased expression and functional activity of V-ATPase were most likely responsible for the decreased in uterine fluid pH observed under progesterone influence.
  5. Lwin OM, Giribabu N, Kilari EK, Salleh N
    J Dermatolog Treat, 2021 Dec;32(8):1039-1048.
    PMID: 32013660 DOI: 10.1080/09546634.2020.1721419
    PURPOSE: This study identifies the potential use of mangiferin gel to promote wound healing in diabetes mellitus (DM).

    MATERIALS AND METHODS: Male rats were rendered diabetes mellitus via intraperitoneal injection of streptozotocin and nicotinamide. Following diabetes development, wound was created at the back of the neck. 1% and 2% mangiferin gel and 1% silver sulphurdiazine (SS) gel (positive control) were applied to the wound for twenty-one (21) days. Fasting blood glucose (FBG) levels were weekly monitored. At the end of the treatment, rats were sacrificed and wound was excised and subjected for histopathological and molecular biological analysis.

    RESULTS: No changes to serum FBG levels was noted throughout the period of mangiferin treatment. Albeit, a significant decrease in the size of the wound with increased in the skin thickness of surrounding the wound were observed. Increased expression and distribution of EGF, FGF, TGF-β, VEGF, PI3K, MMP and Nrf2 and decreased expression and distribution of TNFα and NF-κB p65 were observed in diabetic wound treated with topical mangiferin.

    CONCLUSIONS: Mangiferin has potential to be used as an agent to promote wound healing in diabetic condition.

  6. Dehghan F, Haerian BS, Muniandy S, Yusof A, Dragoo JL, Salleh N
    Scand J Med Sci Sports, 2014 Aug;24(4):e220-9.
    PMID: 24283470 DOI: 10.1111/sms.12149
    Relaxin is a hormone structurally related to insulin and insulin-like growth factor, which exerts its regulatory effect on the musculoskeletal and other systems through binding to its receptor in various tissues, mediated by different signaling pathways. Relaxin alters the properties of cartilage and tendon by activating collagenase. This hormone is also involved in bone remodeling and healing of injured ligaments and skeletal muscle. In this review, we have summarized the literature on the effect of relaxin in musculoskeletal system to provide a broad perspective for future studies in this field.
  7. Khadijah Ramli NS, Giribabu N, Muniandy S, Salleh N
    Theriogenology, 2018 Mar 01;108:354-361.
    PMID: 29294437 DOI: 10.1016/j.theriogenology.2017.12.035
    Precise regulation of vas deferens fluid pH is essential for sperm. However, the mechanisms underlying effect of testosterone on vas deferens fluid pH have never been identified, which could involve changes in expression and functional activity of vacoular (V)-ATPase.

    METHODS: Orchidectomized, adult male Sprague-Dawley rats were treated subcutaneously with 125 μg/kg/day and 250 μg/kg/day testosterone with or without flutamide (androgen receptor blocker) and finasteride (5α-reductase inhibitor) for seven (7) days. Following treatment completion, in vivo perfusion of vas deferens lumen was performed and changes in fluid secretion rate, pH and HCO3- content were measured with and without bafilomycin, a V-ATPase inhibitor. Rats were then sacrificed and vas deferens were harvested and subjected for V-ATPase A1 and B1/2 protein expression and distribution analysis by western blotting and immunohistochemistry, respectively.

    RESULTS: In sham-operated and testosterone-treated orchidectomized rats, higher fluid secretion rate, which was not antagonized by bafilomycin but lower HCO3- content and pH which were antagonized by bafilomycin were observed when compared to orchidectomized-only and orchidectomized, testosterone-treated rats receiving flutamide or finasteride, respectively. Bafilomycin had no effect on fluid secretion rate, HCO3- content and pH in orchidectomized and testosterone-treated orchidectomized rats receiving flutamide and finasteride. V-ATPase A1 and B1/2 proteins were expressed at high levels in vas deferens and were highly distributed at the apical membrane of luminal epithelium and in muscle layer of this organ, mainly in sham and testosterone-treated orchidectomized rats.

    CONCLUSIONS: V-ATPase is involved in acidification of vas deferens fluid under testosterone influence.

  8. Ramli NS, Giribabu N, Muniandy S, Salleh N
    Theriogenology, 2016 Jan 15;85(2):238-46.
    PMID: 26483308 DOI: 10.1016/j.theriogenology.2015.09.036
    Secretions of chloride (Cl(-))- and bicarbonate (HCO3(-))-rich fluid by the seminal vesicles could involve cystic fibrosis transmembrane regulator (CFTR), which activity can be stimulated by cAMP generated from the reaction involving adenylate cyclase (AC). In this study, we investigated levels of CFTR, AC, and cAMP in the seminal vesicles under testosterone influence. Orchidectomized adult male rats received 7-day treatment with 125 or 250 μg/kg/day of testosterone with or without flutamide or finasteride. At the end of the treatment, animals were sacrificed and seminal vesicles were harvested for analyses of CFTR and AC protein expression level by Western blotting. Distribution of CFTR and AC in seminal vesicles was observed by immunohistochemistry. Levels of cAMP and dihydrotestosterone in seminal vesicle homogenates were measured by ELISA. Cystic fibrosis transmembrane regulator, AC, and cAMP levels increased with increasing doses of testosterone (P 
  9. Dehghan F, Muniandy S, Yusof A, Salleh N
    Int J Mol Sci, 2014;15(3):4619-34.
    PMID: 24642882 DOI: 10.3390/ijms15034619
    Ovarian steroids such as estrogen and progesterone have been reported to influence knee laxity. The effect of testosterone, however, remains unknown. This study investigated the effect of testosterone on the knee range of motion (ROM) and the molecular mechanisms that might involve changes in the expression of relaxin receptor isoforms, Rxfp1 and Rxfp2 in the patella tendon and lateral collateral ligament of the female rat knee. Ovariectomized adult female Wistar rats received three days treatment with peanut oil (control), testosterone (125 and 250 μg/kg) and testosterone (125 and 250 μg/kg) plus flutamide, an androgen receptor blocker or finasteride, a 5α-reductase inhibitor. Duplicate groups received similar treatment however in the presence of relaxin (25 ng/kg). A day after the last drug injection, knee passive ROM was measured by using a digital miniature goniometer. Both tendon and ligament were harvested and then analysed for protein and mRNA expression for Rxfp1 and Rxfp2 respectively. Knee passive ROM, Rxfp1 and Rxfp2 expression were significantly reduced following treatment with testosterone. Flutamide or finasteride administration antagonized the testosterone effect. Concomitant administration of testosterone and relaxin did not result in a significant change in knee ROM as compared to testosterone only treatment; however this was significantly increased following flutamide or finasteride addition. Testosterone effect on knee passive ROM is likely mediated via dihydro-testosterone (DHT), and involves downregulation of Rxfp1 and Rxfp2 expression, which may provide the mechanism underlying testosterone-induced decrease in female knee laxity.
  10. Khadijah Ramli NS, Giribabu N, Salleh N
    Steroids, 2018 10;138:117-133.
    PMID: 30003911 DOI: 10.1016/j.steroids.2018.06.012
    Effects of testosterone on expression and functional activity of ENaC, CFTR and NHE in vas deferens were investigated.

    METHODS: Orchidectomized, adult male rats were given 125 and 250 μg/kg/day testosterone subcutaneously, with or without flutamide and finasteride for seven consecutive days. At the end of the treatment, rats were anesthetized and vas deferens were perfused. Changes in vas deferens fluid secretion rate, pH, HCO3-, Cl- and Na+ concentrations were recorded in the presence of amiloride and Cftr inh-172. Rats were then sacrificed and vas deferens were harvested and subjected for molecular biological analysis.

    RESULTS: Testosterone treatment caused the fluid pH and HCO3- concentrations to decrease but secretion rate, Cl- and Na+ concentrations to increase, where upon amiloride administration, the pH and HCO3- concentration increased but Cl- and Na+ concentrations further increased. In testosterone-treated rats, administration of Cftr inh-172 caused all fluid parameters to decrease. In testosterone-treated rats co-administered with flutamide or finasteride, pH and HCO3- concentration increased but fluid secretion rate, Cl- and Na+ concentrations decreased and these parameters were not affected by amiloride or Cftr inh-172 administration. Under testosterone influence, CFTR and γ-ENaC were highly expressed at the apical membrane while NHE-1 and 4 were highly expressed at the basolateral membrane of vas deferens epithelium. Meanwhile, NHE-2 and 3 were highly expressed at the apical membrane.

    CONCLUSIONS: Differential expression of ENaC, CFTR and NHE in vas deferens under testosterone influence indicated the important role of these transporters in creating optimal fluid microenvironment that is essential for preserving male fertility.

  11. Mokhtar HM, Giribabu N, Muniandy S, Salleh N
    Int J Clin Exp Pathol, 2014;7(5):1967-76.
    PMID: 24966906
    Pinopode, a progesterone-dependent endometrial projection which appears during uterine receptivity period, participates in blastocyst implantation. Blastocyst loosely attaches to pinopode via L-selectin ligand (MECA-79). We hypothesized that pinopode and MECA-79 expressions were affected by testosterone. Therefore, the effect of testosterone on pinopode and MECA-79 expressions during uterine receptivity period were investigated.
  12. Mohd Mokhtar H, Giribabu N, Kassim N, Muniandy S, Salleh N
    J Steroid Biochem Mol Biol, 2014 Oct;144 Pt B:361-72.
    PMID: 25125390 DOI: 10.1016/j.jsbmb.2014.08.007
    Estrogen is known to stimulate uterine fluid and Cl(-) secretion via CFTR. This study investigated testosterone effect on these changes in a rat model.
  13. Mokhtar MH, Giribabu N, Salleh N
    In Vivo, 2019 12 29;34(1):225-231.
    PMID: 31882482 DOI: 10.21873/invivo.11764
    BACKGROUND/AIM: It was hypothesized that endometrial tight junction morphology and expression of tight junction proteins i.e., claudin-4 and occludin in the uterus, are affected by testosterone. Therefore, the effects of testosterone on these parameters in the uterus during receptivity period were investigated.

    MATERIALS AND METHODS: Ovariectomized adult female rats were given testosterone (1 mg/kg/day) alone or in combination with flutamide or finasteride between days 6 to 8 of sex-steroid replacement treatment, which was considered the period of uterine receptivity. Ultramorphology of tight junctions was visualized by transmission electron microscopy while distribution and expression of claudin-4 and occludin were examined by immunofluorescence and real-time polymerase chain reaction respectively.

    RESULTS: Administration of testosterone caused loss of tight junction complexity and down-regulated expression of claudin-4 and occludin in the uterus.

    CONCLUSION: Decreased endometrial tight junction complexity and expression of claudin-4 and occludin in the uterus during receptivity period by testosterone may interfere with embryo attachment and subsequent implantation.

  14. Salleh N, Mokhtar HM, Kassim NM, Giribabu N
    J. Membr. Biol., 2015 Dec;248(6):1097-105.
    PMID: 26198330 DOI: 10.1007/s00232-015-9823-8
    Testosterone has been reported to cause a decrease in uterine fluid volume in which this could involve the aquaporins (AQPs). This study aimed to investigate effect of testosterone on uterine AQP-1, 5, and 7 expressions in order to explain the reported reduction in uterine fluid volume under testosterone influence. Ovariectomized adult female rats received peanut oil, testosterone (1 mg/kg/day), estrogen (0.2 µg/kg/day), or combined estrogen plus testosterone for three consecutive days. Other groups received 3 days estrogen followed by 2 days either peanut oil or testosterone with or without flutamide or finasteride. A day after last injection, uteri were harvested, and the levels of AQP-1, 5, and 7 messenger RNA (mRNA) in uterine tissue homogenates were analyzed by real-time PCR (qPCR). Distributions of AQP-1, 5, and 7 proteins in uterus were observed by immunofluorescence. Levels of AQP-1 mRNA were elevated in rats receiving either estrogen or testosterone-only treatment; however, levels of AQP-5 and 7 mRNAs were elevated in rats receiving testosterone-only treatment. In rats pre-treated with estrogen, testosterone treatment resulted in higher AQP-1, 5, and 7 mRNA levels compared to vehicle treatment. Testosterone effects were antagonized by flutamide but not finasteride. Immunofluorescence study showed that AQP-1 was highly distributed in uterine lumenal epithelium following estrogen or testosterone-only treatment. However, AQP-5 and 7 distributions were high in uterine lumenal epithelium following testosterone-only treatment. Testosterone-induced up-regulation of AQP-1, 5, and 7 expressions in uterus could explain the observed reduction in uterine fluid volume as reported under this condition.
  15. Loh SY, Giribabu N, Salleh N
    PeerJ, 2016;4:e2145.
    PMID: 27413634 DOI: 10.7717/peerj.2145
    Testosterone has been reported to cause blood pressure to increase. However mechanisms that underlie the effect of this hormone on this physiological parameter are currently not well understood. The aims of this study were to investigate effects of testosterone on expression of α, β and γ-epithelial sodium channel (ENaC) proteins and messenger RNAs (mRNAs) in kidneys, the channel known to be involved in Na(+) reabsorption, which subsequently can affect the blood pressure. Methods. Adult male Sprague-Dawley (SD) rats were orchidectomized fourteen days prior to receiving seven days treatment with testosterone propionate (125 µg/kg/day or 250 µg/kg/day) with or without flutamide (androgen receptor blocker) or finasteride (5α-reductase inhibitor). Following sacrifice, the kidneys were removed and were subjected for α, β and γ-ENaC protein and mRNA expression analyses by Western blotting and Real-time PCR (qPCR) respectively. The distribution of α, β and γ-ENaC proteins in kidneys were observed by immunofluorescence. Results. The α, β and γ-ENaC proteins and mRNA levels in kidneys were enhanced in rats which received testosterone-only treatment. In these rats, α, β and γ-ENaC proteins were distributed in the distal tubules and collecting ducts of the nephrons. Co-treatment with flutamide or finasteride resulted in the levels of α, β and γ-ENaC proteins and mRNAs in kidneys to decrease. In conclusions, increases in α, β and γ-ENaC protein and mRNA levels in kidneys mainly in the distal tubules and collecting ducts under testosterone influence might lead to enhance Na(+) reabsorption which subsequently might cause an increase in blood pressure.
  16. Mohamed NA, Ramli S, Amin NN, Sulaiman WS, Isahak I, Jamaluddin TZ, et al.
    Med J Malaysia, 2016 04;71(2):62-5.
    PMID: 27326943 MyJurnal
    INTRODUCTION: Nasal colonisation of S. aureus in healthy children was 18% to 30%. One to three percent of them were colonised by Methicillin-resistant Staphlycoccus aureus (MRSA). Although MRSA infection has become increasingly reported, population-based S. aureus and MRSA colonisation estimates are lacking. The main objective of this study was to determine the prevalence of S. aureus carriage among children.

    METHODS: Nasal samples for S. aureus culture were obtained from 250 children from three kindergartens in the Klang Valley, after consent was obtained from the children and their parents. Swabs were transported in Stuart medium, and inoculated on mannitol-salt agar within four hours of collection. Identification and disk diffusion test were done according to guidelines. Polymerase chain reaction was done on MRSA isolates for the presence of mecA and lukS/FPV genes.

    RESULTS: Overall prevalence of S. aureus and MRSA carriage were 19.2% (48/250) and 1.6% (4/250) respectively. mecA gene was present in all isolates, 50% isolates carried Panton-Valentine leucocidin (PVL) gene. Sccmec type I was found in 2 isolates and the remaining isolates has Sccmec type V.

    CONCLUSION: The prevalence of S. aureus and MRSA carriage were similar to other studies. However, risk of contracting severe infection might be higher due to presence of PVL gene in half of the MRSA isolates.
  17. Salleh NM, Tan BA, Arshat H
    Malays J Reprod Health, 1986 Jun;4(1):20-8.
    PMID: 12268569
    PIP: The effectiveness and impact of the Population and Family Health Project in the rural areas of Malaysia is evaluated. A total of 790 women who delivered during the past 3 years from the time of the survey, were identified from 2,013 women aged 15-49 years, during the 2nd Family and Health Survey (1979) in Peninsular Malaysia. The rural health districts which were selected by random sampling include: Perlis, Kubang Pasu, Sebeang Perai Selatan, Kuala Selangor, Hulu Langat, Melaka Utara, Kota Bharu/Tumpat, and Pasir Mas. The pattern of maternal care during pregnancy, delivery and puerperium are examined with respect to selected variables. These women are further classified into 2 groups by type of birth attendant at delivery and these 2 groups are also examined in relation to selected socioeconomic variables. The major proportion of women had their 1st antenatal visit during the 2nd and 3rd trimester of pregnancy. The highest % (28.3%) of 1st antenatal visits occurred during the 5th month of pregnancy. Use of a trained medical practitioner is preferred (82.5%), while only 17.5% of women preferred the services of traditonal birth attendants. Women in this latter group had less education and were in lower income groups, than the former group of women. Majority of women in all ethnic and age groups had no postnatal check after their last childbirth.
  18. Salleh NM, Peng TN, Arshat H
    Malays J Reprod Health, 1986 Dec;4(2):65-71.
    PMID: 12314886
    PIP: Knowledge about contraception was examined in relation to selected socioeconomic variables. A total of 2567 currently married women aged 15-49 years residing in Kuala Lumpur and Petaling Jaya were interviewed. The majority of the women knew of at least 1 contraceptive method. An index termed Contraceptive Knowledge Score (CKS) was used to measure the level of knowledge about contraception. The CKS achieved differed significantly by age, area of residence and ethnic group. The other socioeconomic variables significantly associated with CKS are schooling, occupation, income, childhood residence and age at marriage. These relationships persisted even after adjusting for differences in age, ethnicity and area of residence. Overall the CKS attained have a wide range and there is no significant difference of the mean CKS attained, between users and non-users of contraceptives.
  19. Dehghan F, Muniandy S, Yusof A, Salleh N
    Int J Med Sci, 2014;11(2):180-91.
    PMID: 24465164 DOI: 10.7150/ijms.6283
    The incidence of non-contact knee injury was found higher in female than in male and is related to the phases of the menstrual cycle. This raised the possibility that female sex-steroids are involved in the mechanism underlying this injury via affecting the expression of the receptors for relaxin, a peptide hormone known to modulate ligament laxity. Therefore, this study aims to investigate the effect of sex-steroids on relaxin receptor isoforms (RXFP1 & RXFP2) expression in the ligaments and tendons of the knee.

    METHODS: Ovariectomized adult female WKY rats were treated with different doses of estrogen (0.2, 2, 20 μg/kg), progesterone (4mg) and testosterone (125 & 250μg/kg) for three consecutive days. At the end of the treatment, the animals were sacrificed and the patellar tendon and lateral collateral ligament were harvested for mRNA and protein expression analyses by Real Time PCR and Western blotting respectively.

    RESULTS: RXFP1, the main isoform expressed in these knee structures and RXFP2 showed a dose-dependent increase in expression with estrogen. Progesterone treatment resulted in an increase while testosterone caused a dose-dependent decrease in the mRNA and protein expression of both relaxin receptor isoforms.

    DISCUSSION: Progesterone and high dose estrogen up-regulate while testosterone down-regulates RXFP1 and RXFP2 expression in the patellar tendon and lateral collateral ligament of rat's knee.

    CONCLUSION: Relaxin receptor isoforms up-regulation by progesterone and high dose estrogen could provide the basis for the reported increase in knee laxity while down-regulation of these receptor isoforms by testosterone could explain low incidence of non-contact knee injury in male.

  20. Gholami K, Loh SY, Salleh N, Lam SK, Hoe SZ
    PLoS One, 2017;12(6):e0176368.
    PMID: 28591185 DOI: 10.1371/journal.pone.0176368
    Real-time quantitative PCR (qPCR) is the most reliable and accurate technique for analyses of gene expression. Endogenous reference genes are being used to normalize qPCR data even though their expression may vary under different conditions and in different tissues. Nonetheless, verification of expression of reference genes in selected studied tissue is essential in order to accurately assess the level of expression of target genes of interest. Therefore, in this study, we attempted to examine six commonly used reference genes in order to identify the gene being expressed most constantly under the influence of testosterone in the kidneys and hypothalamus. The reference genes include glyceraldehyde-3-phosphate dehydrogenase (GAPDH), actin beta (ACTB), beta-2 microglobulin (B2m), hypoxanthine phosphoribosyltransferase 1 (HPRT), peptidylprolylisomerase A (Ppia) and hydroxymethylbilane synthase (Hmbs). The cycle threshold (Ct) value for each gene was determined and data obtained were analyzed using the software programs NormFinder, geNorm, BestKeeper, and rank aggregation. Results showed that Hmbs and Ppia genes were the most stably expressed in the hypothalamus. Meanwhile, in kidneys, Hmbs and GAPDH appeared to be the most constant genes. In conclusion, variations in expression levels of reference genes occur in kidneys and hypothalamus under similar conditions; thus, it is important to verify reference gene levels in these tissues prior to commencing any studies.
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