Abstract Estrogen deficiency in women, particularly during menopause, increases bone resorption and decreases bone density, leading to a condition known as osteoporosis. Leucaena leucocephala Lam. de Wit contains flavonols with both estrogenic and anti-estrogenic properties, potentially affecting reproductive and bone health. This research aimed to assess the impact of L. leucocephala leaf extract on blood lipid profiles and the histological structure of lumbar bones in rats subjected to ovariectomy. A total of 28 ovariectomized rats were allocated into four treatment groups: K− (negative control; administered 0.5% CMC-Na), K+ (positive control; given 17β-estradiol at a dose of 0.1 mg/kg body weight/day), P1 (received L. leucocephala extract at 250 mg/kg BW/day), and P2 (received L. leucocephala extract at 300 mg/kg BW/day). All treatments were given daily for 30 consecutive days. Parameters measured included blood lipid levels (total cholesterol, LDL, HDL) and lumbar bone histology, focusing on osteoblast and osteoclast cell numbers as well as trabecular thickness.  The 300 mg/kg BW dose significantly reduced total cholesterol and LDL levels, and increased HDL compared to the negative control. The 250 mg/kg BW dose lowered LDL and total cholesterol; however, did not significantly increase HDL. Both doses increased osteoblast counts and reduced osteoclast counts relative to the negative control, though the changes were not statistically significant. Trabecular thickness decreased in both extract-treated groups compared to the estradiol group.  Administration of L. leucocephala leaf extract at 250 and 300 mg/kg BW/day for 30 days improved blood lipid profiles and modulated lumbar bone histological structure in ovariectomized rats.

Keywords: Bone histology, Leucaena leucocephala, Lipid profile, Osteoporosis, Ovariectomized rats 

Citation:  Wiratmini, N.I., Puspawati, N.M., Setyawati, I., and Pharmawati, M. 2026. Leucaena leucocephala leaf extract modulates lipid profile and bone histology in ovariectomized rats. Natural and Life Sciences Communications. 25(3): e2026064.

Graphical Abstract:

INTRODUCTION

Estrogen deficiency in women due to menopause triggers an increase in bone resorption that exceeds bone formation.  Menopause occurs when ovarian function declines in producing estrogen, resulting in significant calcium loss from the body.  Over time, this leads to reduced bone density, causing bones to become thin, brittle, and porous—a condition known as osteoporosis. In premenopausal women, osteoporosis is generally not observed due to adequate levels of estrogen.  Estrogen, particularly 17β-estradiol (E2) and estrone, are widely recognized as potential inhibitors of bone resorption (Devkota et al., 2012).  In addition to its impact on bone density, postmenopause also rises the chance of cardiovascular disease, primarily due to estrogen deficiency.  According to Yang et al. (2023), estrogen has been shown to lower Low-Density Lipoprotein (LDL) cholesterol, increase High-Density Lipoprotein (HDL) cholesterol, and improve vascular function.

A decline in blood plasma estrogen levels affects bone health in both humans and animals. Dairy cows frequently suffer from hypocalcemia after parturition and from milk fever (parturient paresis), a hypocalcemic disorder associated with the onset of lactation (Goff, 2014). The decrease in plasma estrogen levels after calving leads to enhanced bone resorption by osteoclasts (Devkota et al., 2015).  Laying hens are also vulnerable to osteoporosis due to the high number of eggs produced throughout the laying cycle (Moraes et al., 2020).

Phytoestrogens are substances found in plant that possess properties similar to the hormone estrogen. Due to their structural resemblance to estrogen, they can act as either estrogen agonists or antagonists. In menopausal women, these natural substances are often utilized as substitute for hormone replacement therapy (HRT) to support the body in adjusting to significant hormonal fluctuations and to ease menopausal symptoms.  Lv et al. (2019) reported that supplementation with genistein in hens improved reproductive performance and bone composition during the late laying period.  Studies using ovariectomized rats have shown that phytoestrogens such as quercetin, coumestrol, genistein, and daidzein can reduce bone loss (Leung, 2016).

Leucaena leucocephala Lam. de Wit has been extensively utilized in traditional medicine and as livestock feed.  Species in the Leucaena genus have been reported to contain hydrocyanic acid, leucaenine, epicatechin-3-O-gallate, quercetin-3-O-arabinofuranoside, and apigenin (Aderogba et al., 2010). Showed that in rats with ovariectomies, L. leucocephala leaf extract regulated the estrous cycle and raised uterine weight (Wiratmini et al. 2024) Phytochemical investigation of L. leucocephala leaf extract verified the presence of flavonoids, such as quercetin (Fernandez et al., 2020).  Therefore, it is important to explore the possible impact of L. leucocephala leaf extract on bone tissue structure and blood biochemical profiles in ovariectomized rats.

MATERIALS AND METHODS

Experimental animals and feed

All animal procedures complied with the Animal Ethics Committee, Faculty of Veterinary Medicine, Udayana University (Approval No. B/170/UN14.2.9/PT.01.04/ 2021 dated 19 August 2021). Female Wistar rats (190–200 g) were housed in the laboratory animal facility in plastic cages (33 × 25 × 14 cm) with wire mesh covers and rice husk bedding, under controlled conditions (12-hour light/dark cycle, 25 ± 2°C).  Rats were acclimatized for seven days, given commercial pig concentrate feed (CP 551, PT. Charoen Pokphand Indonesia) and water ad libitum, then monitored through two normal reproductive cycles.

Ovariectomy animal model

Ovariectomy was performed one week after feed adaptation to induce early menopause. Rats were anesthetized intramuscularly with ketamine (50 mg/kg BW) and xylazine (10 mg/kg BW). The procedure was carried out using a ventral incision or transverse abdominal incision technique.

The surgical area was shaved and disinfected with 70% alcohol. A 1 cm transverse incision was made along the midline of the abdomen, slightly to the right, exposing the transverse abdominal muscle.  After incising the muscle, the peritoneal cavity and the fat tissue covering the ovaries became visible.  The ovaries were removed through a single ventral abdominal incision, with both the right and left ovaries excised (Khajuria et al., 2012).  After removal, the peritoneum, muscle, and skin layers were sutured aseptically.  Postoperative care involved treating the surgical wound by applying an antibiotic ointment.

Extract preparation

Leaves of L. leucocephala were collected from plants in the Benoa area, Badung, Bali, Indonesia (8°42'53.8"S, 115°13'21.9"E).  Leaves were detached from the stalk, thoroughly cleaned, dried in the shade to avoid direct sunlight, and subsequently processed into fine powder by grinding, blending, and sieving.

The leaf powder was macerated in 96% ethanol (1:10 w/v) for 72 hours. The extract was then filtered and concentrated under vacuum conditions using a rotary evaporator, resulting in a semi-solid crude form.

Research design and animal treatment

The study used a completely randomized design with four groups of ovariectomized rats: K− (negative control, 2 mL 0.5% CMC-Na), K+ (positive control, estradiol 17β at 0.1 mg/kg BW), P1 (L. leucocephala extract at 250 mg/kg BW), and P2 (L. leucocephala extract at 300 mg/kg BW), with 0.5% CMC-Na as the extract solvent. The sample size in this study was calculated using the formula by Federer (1963): (t-1) (n-1) ≥ 15, where t is the number of treatment groups, and n is the number of subjects per group. To anticipate potential attrition, one additional subject was included in each group, resulting in n = 7 subjects per group.  Treatments began three months post-ovariectomy (Shen et al., 2017) and were administered by oral gavage at 2 mL/rat/day for 30 days.

Blood profile analysis

Rats were fasted for 12 hours prior to sample collection. All animals were anesthetized through intramuscular injection of ketamine–xylazine into the lateral femur. Blood samples (2 mL) were collected from the orbital sinus using microhematocrit tubes. Prior centrifugation for 20 minutes at 4,000 rpm, blood samples were allowed to stand at room temperature for 15 minutes. The resulting serum was used to measure concentrations of Low-Density Lipoprotein (LDL), High-Density Lipoprotein (HDL), and total cholesterol using a spectrophotometric method with a cholesterol assay kit (CHOD-PAP cholesterol method).

Histological sectioning of lumbar vertebra bone

After blood collection, the rats were euthanized, followed by dissection to retrieve the fifth lumbar vertebra.  Bone samples were preserved in neutral buffered formalin (10%) to prepare for histological analysis. The tissue sections were then observed using a light microscope at 400× magnification. The average quantity of osteoblasts and osteoclasts as well as the trabeculae's thickness were assessed.

Identification of active compounds

Liquid Chromatography–Mass Spectrometry (LC-MS) was used for analysis of compounds present in the L. leucocephala leaf extract.  As much as 0.5 grams of the ethanol-based extract were dissolved in 50 mL of methanol (reagent grade). After passing through a 0.22 μm syringe filter, the solution was transferred into a 2 mL vial and put into the LC- the LC-MS instrument for compound identification.

Data analysis

Hematological parameters were evaluated statistically using a two-way ANOVA, performed with SPSS software version 21. To determine significant differences between groups, a post hoc analysis was conducted using Duncan's Multiple Range Test.

RESULTS

Blood serum lipid profile in rats

Table 1 presents total cholesterol, HDL, and LDL levels in rats treated with L. leucocephala leaf extract for 30 days. ANOVA revealed significant differences (P < 0.05) among groups, with post hoc analysis showing the highest cholesterol in K− and the lowest in K+. P2 did not differ from K+, whereas P1 was significantly higher, indicating that 300 mg/kg BW extract effectively reduced cholesterol.

For HDL, K+ showed the highest levels, significantly different from K−, P1, and P2. No difference was observed between K− and P1, while P2 differed significantly, suggesting that 17β-estradiol and 250 mg/kg BW extract produced the best HDL outcomes.  LDL analysis showed no significant differences among K+, P1, and P2. However, both extract doses (250 and 300 mg/kg BW) reduced LDL compared to K−. Data are summarized in Table 1.

Table 1. Mean of total cholesterol, HDL, and LDL in the blood serum of ovariectomized female rats treated with L. leucocephala leaf extract for 30 days.

Note: Values are presented as mean ± SD. Different letters within the same column indicate statistically significant differences (P < 0.05). Treatments: K− (2 mL of 0.5% CMC-Na/day); K+ (Estradiol 17 β at 0.1 mg/kg BW/day); P1 (250 mg/kg BW/day of L. leucocephala leaf extract); P2 (300 mg/kg BW/day of L. leucocephala leaf extract).

Osteoblast and osteoclast cell counts and trabecular bone thickness

Analysis of lumbar vertebral tissue under the microscope demonstrated that the average number of osteoblast cells differed significantly among the K−, K+, and P2 groups, but did not differ significantly from the P1 group (Table 2, Figure 1). Meanwhile, the average number of osteoclast cells did not show any significant distinctions between the K− and P1 groups, nor between the K+ and P2 groups (Table 2, Figure 2).

Measurement of trabecular thickness indicated that the K− group had the thinnest trabeculae, which exhibited notable differences in contrast to other treatment groups. There was no variation was detected in trabecular thickness between the P1 and P2 groups (Table 2, Figure 3 and 4).

Table 2. Mean of total cholesterol, HDL, and LDL in the blood serum of ovariectomized   female rats treated with L. leucocephala Leaf extract for 30 days.

Note: Values are presented as mean ± SD. Different letters in the same column indicate significant differences (P < 0.05). Treatment: K− (2 ml of 0.5% CMC-Na/day); K+ (Estradiol 17 ß at 0.1 mg/kg BW/day); P1 (250 mg/kg BW/day of L. leucocephala leaf extract); P2 (300 mg/kg BW/day of L. leucocephala leaf extract).

Figure 1. Histological section of lumbar bone stained with hematoxylin-eosin (HE) (400× magnification). K− (rats administered 0.5% CMC-Na); K+ (rats treated with estradiol 17β at a dose of 0.1 mg/kg BW/day); P1 (rats received L. leucocephala leaf extract at a dose of 250 mg/kg BW/day); P2 (rats given L. leucocephala leaf extract at a dose of 300 mg/kg BW/day). The abbreviation bm refers to bone marrow, and white arrows indicate osteoblast cells.

Figure 2. Histological section of lumbar bone stained with hematoxylin-eosin (HE) (400× magnification). K− (rats administered 0.5% CMC-Na); K+ (rats treated with estradiol 17β at a dose of 0.1 mg/kg BW/day), P1 (rats receiving L. leucocephala leaf extract at a dose of 250 mg/kg BW/day), and P2 (rats given L. leucocephala leaf extract at a dose of 300 mg/kg BW/day). Red circles indicate osteoclast cells.

Figure 3. Histological section of lumbar bone stained with hematoxylin-eosin (HE) (400× magnification). K− (rats administered 0.5% CMC-Na); K+ (rats treated with estradiol 17β at a dose of 0.1 mg/kg BW/day); P1 (rats receiving L. leucocephala leaf extract at a dose of 250 mg/kg BW/day); and P2 (rats administered L. leucocephala leaf extract at a dose of 300 mg/kg BW/day). The abbreviations tb, vc, and bm refer to trabecular bone, vascular cavity, and bone marrow, respectively.

Figure 4. Histological section of lumbar bone stained with hematoxylin-eosin (HE) (100× magnification). K− (rats administered 0.5% CMC-Na); K+ (rats treated with estradiol 17β at a dose of 0.1 mg/kg BW/day); P1 (rats receiving L. leucocephala leaf extract at a dose of 250 mg/kg BW/day), and P2 (rats given L. leucocephala leaf extract at a dose of 300 mg/kg BW/day). The abbreviation P refers to the periosteum, and white arrows indicate areas of bone erosion.

To evaluate the potential toxic effects of the treatment, it is important to record the body weight of the test animals at the start and end of the course of treatment, as well as monitor daily food intake. Experimental results suggested that administering L. leucocephala leaf extract did not inhibit growth. This was supported by statistical analysis showing that no notable variations in body weight or dietary intake across treatment and control groups. Rats body weight, both before treatment (initial weight) and after treatment (final weight), showed no significant differences. Similarly, food intake did not differ significantly among all groups, both control and treatment (Table 3).

Table 3. Average initial weight, final weight, and feed intake of female rats administered L. leucocephala leaf extract for 30 days.

Note: Values are presented as mean ± standard deviation (SD). Different letters within the same column indicate statistically significant differences (P < 0.05). Treatments: K− (2 mL of 0.5% CMC-Na/day); K+ (Estradiol 17β at a dose of 0.1 mg/kg BW/day); P1 (L. leucocephala leaf extract at a dose of 250 mg/kg BW/day); P2 (L. leucocephala leaf extract at a dose of 300 mg/kg BW/day).

Qualitative phytochemical determination of L. leucocephala leaf extract

Phytochemical analysis of L. leucocephala leaf extract using LC-MS spectra identified the presence of flavonoids, polyphenols, lignans, and steroids, which function as phytoestrogens (Figure 5 and Table 4).

Figure 5. LC-MS chromatogram of L. leucocephala leaf extract.

Table 4. Identification of potential phytoestrogen peaks in L. leucocephala leaf   extract as determined by LC-MS.

DISCUSSION

Body weight and feed intake are important basic parameters that can be used to assess the toxicity of test substances. Toxic compounds in certain materials may interfere with digestive enzyme function, resulting in a notable reduction in the appetite of rats. This, in turn, can negatively affect their development (Karnam et al., 2014). In the present study, neither ovariectomy nor oral administration of estradiol or L. leucocephala leaf extract affected the body weight or feed consumption of the rats.

The ovariectomized rats used in this study served as a model for postmenopausal women. During menopause, estrogen secretion decreases due to ovarian atrophy. This decline in estrogen levels can lead to increased total cholesterol, elevated LDL, and reduced HDL levels.

The blood lipid profile parameters of rats treated with L. leucocephala leaf extract in this study showed that the K− group displayed the lowest HDL levels, while exhibiting the highest levels of total cholesterol and LDL compared to other treatment groups. This outcome is attributed to the reduction of estrogen following ovariectomy. The increase in HDL and decrease in LDL observed in the K+ group resulted from the administration of Estradiol 17β, a synthetic estrogen. Similarly, the improvement in HDL and reduction in LDL levels in the P1 and P2 groups were due to the administration of L. leucocephala leaf extract over a 30-days.

In this study, L. leucocephala leaf extract was found to contain flavonoids, lignans, phenolics, phenolic acids, and steroids. According to Dixon (2003), the main classes of phytoestrogens are lignans and isoflavones. Lignans are another class of phytoestrogens commonly found in grains, legumes, coffee and tea, cocoa, flaxseeds, and certain fruits (Durazzo et al., 2018). Lignans exhibit estrogenic properties because their two phenolic rings mimic the A and D rings of steroid hormones, enabling them to bind to estrogen receptors (ER). Lignans display varying degrees of estrogenic activity, and their estrogenic potency contributes to their diverse biological functions (Kiyama, 2016).

Flavonoids can inhibit Apolipoprotein B (ApoB) secretion and enhance LDL receptor expression, thereby increasing the uptake of LDL cholesterol (Casaschi et al. 2002). In addition, Kim et al. (2014) reported that rats fed soy milk containing phytoestrogens, specifically daidzein and genistein, experienced a reduction in LDL levels due to increased expression of LDL receptors.

According to Keika and Veisian (2021), serum LDL and triglyceride levels tend to be higher in postmenopausal women with reduced bone density. Animal studies have shown that a high-cholesterol diet can lead to the production of lipid oxidation products that interfere with bone regeneration, reduce bone mineralization, and impair bone mechanical strength (Pirih et al., 2012).

The histological structure of the lumbar bone in rats treated with L. leucocephala leaf extract showed significant improvement compared to the K− group.  This improvement is attributed to the presence of secondary metabolites, particularly flavonoids such as quercetin and kaempferol, which are classified as phytoestrogens.  According to Oh and Chung (2004), isorhamnetin and kaempferol are phytoestrogens that function as Selective Estrogen Receptor Modulators (SERMs) and can be used in hormone replacement therapy (HRT) for postmenopausal women. Similarly, Hassan et al. (2013) reported that L. leucocephala leaf extract contains phytoestrogens in the form of flavonoids such as caffeic acid, isorhamnetin, chrysoeriol, isorhamnetin 3-O-galactoside, and kaempferol-3-O-rutinoside. A study by Andini et al. (2023) demonstrated that the flavonoid quercetin could be used as a therapeutic agent for menopausal symptoms in vaginal tissue. Quercetin has the ability to interact with estrogen receptors (ER), modulating the expression of genes regulated by estrogen, and exhibiting estrogen agonist activity.

Another group of compounds identified in L. leucocephala leaf extract in this study is polyphenols. According to Chadva et al. (2024), phytoestrogens are natural non-steroidal polyphenolic compounds found in plants that can mimic the action of estrogen.  Estrogen is essential for bone maintenance as it helps regulate the equilibrium between bone-forming osteoblasts and bone-resorbing osteoclasts.  In this study, the K− group exhibited the lowest number of osteoblast cells. This is likely due to estrogen deficiency caused by ovariectomy. Wanderman et al. (2018) found that ovariectomy in rabbits induced significant trabecular bone loss within 18 weeks. Similarly, Dawson-Hughes (1996) reported that reduced estrogen levels following ovariectomy—a condition that models postmenopausal women—leads to rapid bone mineral loss, reduced bone mass, and increased bone resorption, ultimately resulting in osteoporosis.  Estrogen deficiency disrupts the equilibrium between bone-resorbing osteoclasts and bone-forming osteoblasts.

A similar condition was observed in the P1 treatment group. Administration of L. leucocephala leaf extract at 250 mg/kg body weight/day for 30 days did not increase the number of osteoblast cells. Although Fernandes et al. (2020) reported that administering L. leucocephala extract at the same dose to ovariectomized rats was able to increase estrogen levels in the blood, the increase in estrogen concentration at this dose may not have been sufficient to activate osteoblast cells.

In the K+ and P2 treatment groups, there was a rise in the count of osteoblast cells, indicating that administration of synthetic estrogen in the K+ group and L. leucocephala leaf extract at a dose of 300 mg/kg body weight/day for 30 days was able to stimulate osteoblast proliferation. The increase in osteoblast count observed in the P2 group can be attributed to the phytoestrogen content of L. leucocephala extract, which raises endogenous estrogen levels sufficient to activate osteoblast differentiation.  This interpretation is further supported by findings from Kantawong
et al. (2022), who demonstrated that Cuscuta japonica extract was able to stimulate osteoblast differentiation by upregulating osteogenic markers such as OPN (Osteopontin), OCN (Osteocalcin), and type I collagen.

Estrogen is essential for the activation of osteoblasts within the endosteal tissue. The role of osteoblasts is to synthesize the organic components of the bone matrix. When active, these cells exhibit a cuboidal shape (Figure 4.2, K+ and P2), whereas in their inactive state, they appear flattened (Figure 4.2, K0 and P1).  Among the phenolic constituents identified in L. leucocephala extract is cinnamic acid.  It was reported that treatment with cinnamic acid promoted osteoblast differentiation by enhancing the expression of osteogenic markers and led to an increase in bone density (Hong et al., 2022).

The mean number of osteoclasts was highest in the K0 and P1 groups compared to K+ and P2. This indicates that the 250 mg/kg body weight/day dose of L. leucocephala extract did not effectively reduce osteoclast numbers. Osteoclasts (Figure 4.3), are both multinucleated and large, and are responsible for bone mineral and matrix resorption by secreting proteolytic enzymes (Figure 4.2, K0 and 4.3). In contrast, the average number of osteoclasts in the K+ and P2 groups decreased, which may be attributed to the elevated estrogen levels induced by the phytoestrogens present in the 300 mg/kg extract, sufficient to inhibit osteoclast differentiation.  Nepetoidin B, a phenolic compound was identified in this study.  A study by Jang et al. (2020) found that nepetoidin B had anti-osteoclastogenic activities thus inhibit osteoclast differentiation.

Osteoclasts are derived from hematopoietic stem cells and form resorption pits by secreting protons to dissolve hydroxyapatite and releasing cathepsin K to break down type I collagen. Excessive bone resorption is often caused by increased osteoclast activity that outpaces osteoblast-mediated bone formation (Arnett, 2013). Estrogen exerts both direct and indirect effects on osteoclasts. Indirectly, it influences osteoclast differentiation, activation, and apoptosis. In terms of differentiation and activation, estrogen inhibits the expression of receptor activator of nuclear factor-kappa B ligand (RANK-L) and macrophage colony-stimulating factor (M-CSF) from osteoblastic stromal cells, preventing the RANK-L and RANK interaction through increased osteoprotegerin (OPG) production. Moreover, estrogen indirectly suppresses cytokines that promote osteoclastogenesis, such as IL-6, IL-1, and TNF-α, while simultaneously stimulating osteoblasts to produce TGF-β, which induces earlier apoptosis of osteoclasts. Directly, estrogen binds to estrogen receptors on osteoclasts, suppressing precursor differentiation and mature osteoclast activation (Liu and Zhang, 2015). 

In this study, trabecular thickness measurements showed that the K0 group (ovariectomized rats) exhibited the thinnest trabeculae (Figure 1), which differed significantly from the other groups. This indicates that reduced estrogen levels due to ovariectomy lead to decreased trabecular thickness. Trabecular bone plays a critical role in overall bone strength (Launey et al., 2010). These findings are consistent with the study by Khajuria et al. (2015), in which histopathological analysis of bones from ovariectomized rats revealed a reduction in the number and thickness of intratrabecular spaces and trabecular widening, indicating bone loss induced by impaired bone formation and enhanced bone resorption.

The finding of this study revealed no notable variation in trabecular thickness between the K+ group and the P1 and P2 treatment groups. This suggests that trabecular thickness in rats treated with Estradiol 17β was comparable to that in rats treated with L. leucocephala leaf extract at doses of 250 and 300 mg/kg body weight/day.

The primary mechanism through which phytoestrogens such as flavonoids benefit bone health is associated with their estrogenic activity. Phytoestrogens such as soy isoflavones, lignans, and coumestrol possess weak affinity for estrogen receptors, particularly ERβ, and have been extensively studied in cell culture systems for their estrogen-like characteristics (Tang et al., 2011). According to Janas et al. (2020), phytoestrogen consumption may help prevent bone fractures by increasing the inorganic content of bone and collagen fiber composition. Phytoestrogens are also known to influence bone resorption by reducing bone degradation through inhibition of ALP (alkaline phosphatase), osteocalcin, and C-terminal cross-linked telopeptide of type I collagen (CTX-1). Additionally, they may prevent trabecular thinning caused by estradiol deficiency and reduce osteoclast numbers in the tibial metaphysis. CTX-1 is a bone resorption marker produced by osteoclast activity. In another study, approximately 39 phytoestrogen compounds were predicted to have anti-osteoporotic activity through an in-silico approach using the Raloxifene web tool and Swiss ADME (Chavda et al., 2024).

CONCLUSION

Administration of L. leucocephala leaf extract at doses of 250 and 300 mg/kg BW/day for 30 days was effective in improving blood lipid profiles and the histological structure of the lumbar bone in ovariectomized rats.

AUTHOR CONTRIBUTIONS

Ngurah Intan Wiratmini: Conceptualization (Lead), Methodology (Lead), Validation (Lead), Formal Analysis (Lead), Investigation (Equal), Resources (Equal), Writing Original Draft (Lead), Writing-Review & Editing (Equal), Supervision (Lead), Project Administration (Equal); Ni Made Puspawati: Formal Analysis (Equal), Data Curation (Equal), Investigation (Equal); Iriani Setyawati: Investigation (Equal), Writing Original Draft (Lead), Writing-Review & Editing (Equal), Data Curation (Equal); Made Pharmawati: Investigation (Equal), Writing Original Draft (Lead), Writing-Review & Editing (Lead), Resources (Supporting).

CONFLICT OF INTEREST

The authors declare that they have no conflicts of interest.

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