Obesity has become a major problem which lead to many chronic diseases such as insulin resistance, type 2 diabetes, cardiovascular disease and polycystic ovary syndrome (Guilherme et al., 2008). The over consumption of high calory diet which lead to reduction in energy utilization and subsequently results in insulin resistance promote the formation of obesity (X.A. et al., 2013). The high fat diet not only shows sign of metabolic impairment it also disturbs the female reproductive performance which can lead to polycystic ovary syndrome (PCOS). Polycystic ovary syndrome is the most common endocrinopathy and most frequent cause of infertility in women reproductive age. It is characterize by hirsutism, acne, anovulation, menstrual irregularities, hypeandrogenemia, hyperinsulinimea and polycystic ovary (Franks, 1995; Goodarzi et al., 2011). According to clinical diagnosis, PCOS requires the presentation of two of the three symptoms of hyperandrogenism, chronic anovulation and polycystic ovaries (Group, 2004). However, PCOS involve many metabolic impairments, the important role in pathophysiology of PCOS is 50–70% of patients have insulin resistance (Dunaif et al., 1989). In much severe cases, PCOS will in time develop to musculoskeletal disorders such as osteoporosis (Katulski et al., 2014), osteoarthritis (Sanchez-Santos et al., 2018) and tendinitis (Jewson et al., 2018). Due to the metabolic impairment in PCOS, it can lead to impairment skin cutaneous leading to loss of elasticity. The inflammatory plays a major role in the development of PCOS and other metabolic problem.
Aqueous extracts of Labisa pumila (LP) is a herbal formula used in this study to determine the efficacy of its natural compound in correcting the effects of osteoporosis, hormonal imbalances and cutaneous disorders. In this study, we presented results showing that orally administrated different concentration of LP25 and LP50 (as stated in Methodology) decreases the osteoporosis, hormonal imbalances, inflammatory mediators level and increases the skin elasticity without any distinct toxicities in PCOS rats induced by HFD.
This study used two doses of LP alone (25 mg/kg and 50 mg/kg) were given at 5 times a week for a duration of 90 days duration of treatments to evaluate the dose effects on modulation of c-reactive protein, TNF-alpha, insulin, serum estradiol and testosterone levels pertaining to the osteoporosis effects by ELISA assay. It is important to note LP extract has been shown to be safe with an LD50 of more than 5.0 g/kg (M.F. et al., 2007). According to other studies on LP extract was reported to exhibit no-adverse-effect-level (NOAEL) at the dose of 50 mg/kg in sub-acute (G.D. et al., 2009). At dose 1000 mg/kg in sub-chronic (S.C, 2004) and 800 mg/kg was reported to show no adverse effect level (NOEL) in reproductive toxicity studies (M.F. et al., 2007). However, in human, the effective doses normally taken by women are around 500–1000 mg/kg daily. Which is similar to 25 mg/kg and 50 mg/kg when calculate for animal trial. Therefore, the doses used in this study are considered to be safe. From the study, it showed that significant effects of anti-osteoporosis was detected in this experiment of PCOS rats treated with LP alone.
Labisia pumila contained phytoestrogenic properties that can upregulate the synthesis of estrogen in human. The antioxidant effects initiated by the estrogen can decrease lipid peroxidation of that will result in oxidative damage and indirectly help in free radical scavenging (S.P. et al., 2002). Furthermore, estrogen deficiency result in the decline bone formation and in turn increase bone resorption process leading to osteoporosis. It can be said that osteoporosis is the systemic skeletal disease characterized by low bone mass and micro-architecture deterioration of bone tissues which results in increase bone fragility and fracture (Norhayati et al., 2014). It is important to understand estrogen is a hormone that has a profound effect on bone physiology. It keep the bone turnover rate at balance and reduces the cause of osteoporosis which can be seen postmenopausal women.
Therefore, the doses used in this study are considered to be safe. From the study, it showed that significant effects of anti-osteoporosis were detected in-group of PCOS rats treated with LP25 and LP50. This indicates that extracts of LP alone managed to reduce the level of C-reactive protein, a cytokine that is used as an indicator for bone mass density. Increase levels of c-reactive protein in blood indicates a detrimental effects of this inflammatory cytokines towards bone health and mass, thus contributing towards the risk of fractures (Huang and Schooling, 2017).
In this study, it was found that the treatment of PCOS rats with LP25 and LP50 alone significantly reduced the levels of TNF-α while the level in normal untreated and placebo groups of rats showed an increase in TNF-α level at a significant rate. It was noted that increase in the level of TNF-α correlates with other parameters that were investigated in this study and this point towards the outcome following PCOS induction in rats in term of bone calcium contents and micro-computed topography (micro-CT) results. TNF-ɑ is a bone resorbing cytokine that promotes bone resorption by activating the mature osteoclast or by stimulating the proliferation and differentiation of osteoclast (Lerner and Ohlin, 1993). The finding were consistent with fathilah et al 2012 and Choi et al 2010 that LP extract inhibit the TNF-α production in the experimental study (Choi et al., 2010; Fathilah et al., 2013). TNF-α promotes osteoclastogenesis by activating NF-κB transcription factor, which is also induced by RANK/RANK-L (Choi et al., 2010; Gilbert et al., 2005). Activation of NF-κB is a key target for TNF-α action through TNF receptor- 1 (TNFR-1), which is expressed on macrophages (Lam et al., 2000). This promotes pro-osteoclasts maturation even in the absence of RANK/RANK-L signaling (Bailey and Etherington, 1980). Tumor necrosis factors-alpha also enhances osteoclast differentiation by increasing the expression of M-CSF and RANK-L in osteoblasts (Affinito et al., 1999).
In this study, it was noted that the levels of estradiol was depleting in the placebo group while the treated group showed a normal levels of estradiol. At the same time, the levels of testosterone was at the normal level and elevated in the place group while the testosterone levels in treated group was at the normal range. The study shows that the LP extract proven to successfully replace the loss of estrogen due reduction in placebo group. In women with PCOS, secretion rate and metabolism of androgens and estrogens are impaired and androgens levels increase (Abasian et al., 2018). Estrogen plays a key role in the development and maintenance of appropriate bone mass. Estrogen regulates the adipocyte differentiation as well as skeletal growth and bone homeostasis. Estrogen acted on osteoblast and osteoclast for bone formation. However, in PCOS the estrogen and androgen were greatly altered which probably has great importance for bone metabolism in women. Estrogen deficiency also can be linked to the increase in food intake and body weight gain which result in obesity. Thus estrogen deficiency increase in adipose tissue proliferation on visceral fats which results in many chronic disease such as cardiovascular disease, osteoporosis and obesity. The LP extract able replace the hormone leptine resistin and adiponectin which in deficient placebo group. Which show the treatment group LP 25 and LP 50 able to maintain normal body weight gain.
The LP extract able to increase the insulin sensitivity of treated group LP 25 and LP 50. The treatment of LP 25 and LP 50 does not affect the body weight gain of PCOS rats. It is reported that PCOS are more insulin resistant and body weight gain (Dunaif et al., 1989). This may be due to the phytoestrogenic effect of LP extract. According Manneras et al 2010 the supplementation of LP extract increases the insulin sensitivity up to 36% as well as improving the lipid profile of PCOS rats without affecting the body weight composition (Mannerås et al., 2010). LP extract possess uterotropic effects which regulate the body weight and lipid proliferation. Previously reported that reduction in plasma resistin increases the insulin sensitivity (26). This is linked to the high level adiponectin in PCOS rat treated with LP extracts (Weyer et al., 2001). Adiponectin is closely related to the metabolic abnormalities such as insulin sensitivity and obesity.
Results from bone scanning electron microscopy, micro-computed topography (micro-CT) and bone calcium contents showed that the administration of LP to the rats with PCOS have successfully replaced the estrogen from defective ovary in these PCOS rats at dosage LP 25 mg/kg and LP 50 mg/kg. Phystoestrogen properties in LP can have an intricate pathways of action in improving the metabolic disorders such as PCOS and osteoporosis. It is reported that PCOS is a linked to ovulatory dysfunction, hyperandrogenism, abdominal fat, insulin resistance and polycystic ovaries (Group, 2004). In a much severe cases. PCOS can develop the loss of bone mass and density which can lead to osteoporosis. Osteoporosis is defined as systemic skeletal disease accompanied by low bone mass and microarchitecture deterioration of bone tissue. This consequently increase the chances to bone fragility and fractures (Grant et al., 1996). However, World Health Organization (WHO) reported osteoporosis can happened when the bone mineral density falls more than 2.5 Standard Deviations (SD) which is below the standard reference of young femaler maximum bone mineral density (Kanis et al., 2008). Form this experiment, LP 25 and LP 50 manage to significantly reduce the osteoporosis inflammatory cytokine TNF-ɑ and C-reactive protein which responsible in the bone formation and bone resorption. Supplementation of LP extract provide a beneficial aspect to estrogen deficiency women. It is reported at the age of 35–40, the bone mass in females begin to decline slowly, followed by a dramatic bone loss after menopause due to estrogen deficiency or surgical ovariectomy.
The result from skin and vaginal elasticity through biochemical testing did not altered the increase in elasticity of both tissues. However, the elasticity is still in normal range suggesting that LP extract did not change or possess any form of elasticity. The elasticity can be speculated whether to increase the concentration of LP extract given to PCOS rats. Other study suggested that the supplementation of LP extract can upregulate the synthesis of collagen human fibroblast cells (Choi et al., 2010). The LP extract can be used to protect the human skin from reactive oxygen species expose from UVB rays. Due to antioxidant and phenolic compound such as quercetin. This compound possesses rejuvenating effects on the skin repairing.