1. Introduction

ne of the endocrine disorders of women is known as polycystic ovary (PCO) which is characterized by polycystic feature of ovary along with the hyperandrogenism and ovulatory dysfunction (1,2). Also, one who suffers from PCO may show a higher exhibition of pro-inflammatory agents such as nitric oxide (NO) (3).

We have already shown that the chronic use of L-arginine, a precursor of NO, in rats with diestrous phase may induce the PCO alongside with lipid metabolism malfunction (3), the characteristics that mark the PCO syndrome (PCOS).

Since, by reviewing of literature, the uteri of those suffering from PCO have not been much studied, we sought to evidence pathologically the PCO model’s uterus. In this work, the female Wistar rats were treated chronically with Larginine to provide the PCO model. Also, the experimental animals’ uteri were examined to provide valuable data representing the involvement of NO in pathophysiology of the disorder.

2. Materials and Methods

2.1. Animals

Wistar rats (body weight 200-250 g) were purchased from Pasteur Institute of Iran and were retained under standard conditions (21 ± 3°C and 12-h light/dark cycle) with food and water ad libitum. All experiments were approved by local Ethical committee.

2.2. Drugs

L-arginine (Merck Co., Germany) was injected i.p. for a 9-day period/ once per day. The vehicle (saline at 1 ml/kg, i.p.) was used in control group.

2.3. Female Cycle Test

Since female rats with 4-5 day sexual cycle are always in diestrous (4,5) unless in case of having mating with the male rat, so, the rats were kept virgin in the present study to avoid the change in female sexual cycle.

2.4. Drug administrations

Animals were randomly divided into the L-arginine (50 mg/kg), and saline control (1 ml/kg) groups (n=6). They were injected the agent or saline intraperitoneally (i.p.) once a day during the 9 days period.

2.5. Surgery procedure

The treatment groups were anesthetized by an overdose of diethyl ether. Then, a midline incision in the lower abdomen area was performed. The ovaries and uteri were biometrically examined and dissected out. They were collected in 10% formalin for histological examination.

2.6. Histological investigation

The collected tissues were processed and sectioned at a thickness of 3-4 µm. They were stained by Hematoxylin and Eosin (H&E) method (6). The thin sections were then dehydrated, cleared, and eventually mounted with entellane (Merck Co., Germany) and coverslipped. The prepared slides were evaluated with light microscope (Olympus, Japan) at 4-40X.

2.7. Image analysis

The photomicrographs were assessed in areas of 100-µm² with an aid of Image Tool program (UTHSCSA, version 2.03), the free image processing and analysis program for Microsoft Windows.

2.8. Statistical analysis

All data were first assessed by Kolmogorov-Smirnov (K-S) to show the equality to analysis by variance (ANOVA). The ANOVA was then performed using SPSS software (version 13.0; SPSS, Inc., Chicago, IL), followed by post-hoc test. Statistical significance was considered at p< 0.05. All data are expressed as Means ± SEM. The photos were examined in an area of 100-µm² using the Image Tool program.

3. Results

3.1. Histology

The ovaries from the L-arginine-treated group (50 mg/kg, chronically) showed cystic formations (Fig. 1B) as compared to control samples (Fig. 1A), the aspects certifying the polycystic ovary (PCO) example.

The uteri samples of those received chronically L-arginine (50 mg/kg) (Fig. 1C) presented pathological evidence to those belonged to the controls (Fig. 1D). Due to activation of the L-arginine-related metabolic pathway, the uterus wall revealed the aspects of swelling, the proliferation and angiogenesis, suggesting an inflammatory process involvement.

3.2. Biometrical value (diameters of uteri)

The uteri diameters were calculated in all groups. They showed changes in L-arginine-treated group as compared to those obtained from saline group. The uteri of rats treated with L-arginine showed a significant increase when compared with the control group (p

4. Discussion

This research study showed that chronic treatment of rats with a nitric oxide (NO) agent, L-arginine, induces the polycystic ovary (PCO) formation as well as the uteri inflammation.

The pro-inflammatory NO participates in endocrine physiological and pathophysiological events (7). The enzyme nitric oxide synthase (NOS) produces the NO by the oxidation of terminal guanidino nitrogen of arginine (7). We have already demonstrated the NOS activation with NADPH-diaphorase (8). The molecule NO is well introduced as a local inflammatory generator (9). The present work also provides support for a functional role of the NO in the ovarian and uterine events. This plan further supports our previous study (8), denoting that

Figure 1. Photomicrographs of ovaries and uteri from control (A, C), and L-arginine-treated (B, D) rats.