Effects Of Diabetes Mellitus On Sexual Function, Histomorphology And α-smooth Muscle Actin Expression in Cavernous Body Of Penis in Rats
Mar 05, 2025
Abstract:
Objective: To observe the sexual function of diabetes model rats and explore the possible mechanism of secondary sexual dysfunction caused by hyperglycemia.
Methods The experimental rats were randomly divided into the blank group (n=5) and the diabetes group (n=15). After the diabetes animal model was successfully constructed, the sexual function of two groups was evaluated. The histological characteristics of testis in two groups were observed. The expression of α-smooth muscle actin (α-SMA) in the penile tissue of two groups was measured. Results The levels of serum sex hormones were significantly decreased (P<0.01). Histomorphological observation showed that in the diabetes group the interstitial tissue of testis was disordered; Compared with the blank group, the number of smooth muscle cells in the penis tissue was reduced, and the degree of fibrosis was aggravated. The expression of α-SMA positive cells in the diabetes group was significantly decreased (P<0.01).
Conclusion The decreased levels of sex hormones, the decreased number of smooth muscle cells in cavernous body of penis, the decreased expression of α-SMA in cavernous body of penis, the disordered structure of interstitial tissue of testis, the morphological changes of smooth muscle cells and the aggravation of fibrosis in the diabetes rats are important phenotypic features of this disease. Exploring the diagnosis and treatment of this disease with this phenotypic features will be of great significance.
Keywords: diabetes mellitus; erectile dysfunction; sexual function; sex hormone; α- smooth muscle actin expression
Herbal Cistanche Supplements For Erectile Dysfunction Caused By Diabetes
Diabetes mellitus (DM) has long been a focal point of attention both domestically and internationally due to its high incidence and prevalence [1]. The various complications induced by DM significantly impact the quality of life of patients. Among these, the incidence of erectile dysfunction (ED) in male DM patients is markedly elevated. Currently, DM has become a high-risk group for ED [2–3]. Therefore, conducting in-depth studies on DM-induced ED (DMED) holds great significance. This study aims to observe the effects of DM on erectile function, sex hormones, the expression levels of α-smooth muscle actin (α-SMA) in the penile cavernous body, and the morphology of testicular interstitial and penile cavernous smooth muscle tissues in rats, preliminarily exploring the potential pathological basis of ED induced by DM.
1 Materials and Methods
1.1 Experimental Animals and Reagents
Twenty adult SPF-grade male SD rats with uniform body weight were selected. The main experimental reagents included streptozotocin (STZ) and apomorphine (APO), as well as antibodies required for detecting α-SMA expression. This study was approved by the Animal Welfare and Ethics Committee of Dongzhimen Hospital, Beijing University of Chinese Medicine (No. 17-04).
1.2 Experimental Methods
1.2.1 Establishing a Diabetic Rat Model
Twenty experimental SD rats were randomly divided into a control group (5 rats) and a diabetes group (15 rats). STZ was dissolved in citrate buffer to prepare a 10 mg/mL solution. The diabetes group was intraperitoneally injected with STZ solution at a dose of 55 mg/kg to induce the DM model rats. Blood glucose levels >16.7 mmol/L at any time after 5 days of STZ intraperitoneal injection were considered the standard for successful DM model establishment [4].
1.2.2 Erectile Function Test in Rats
After confirming successful DM modeling, the rats were weighed and placed in a transparent glass box. The APO solvent (100 μg/kg) was subcutaneously injected at the back of the neck based on body weight. The surrounding environment was kept quiet, and the indoor lighting was adjusted to a level just sufficient for observing penile erection in rats. The number of penile erections within 30 minutes after subcutaneous injection was observed and recorded. Criteria for penile erection: penile enlargement, retraction of the foreskin, exposure of the glans accompanied by hyperemia and redness, and visibility of the middle and lower segments of the penis.
1.2.3 Measurement of Sex Hormones in Rats
The serum sex hormone levels of the two groups were measured using the radioimmunoassay method, following the specific procedures outlined in the radioimmunoassay kit instructions.
1.2.4 Histopathological Observation of Testicular Interstitial Tissue
After collecting blood from the abdominal aorta of the rats, the right testicle was uniformly extracted. The testicle was washed with PBS buffer and fully fixed in paraformaldehyde for 72 hours. The tissue was embedded in paraffin, sectioned, stained with hematoxylin-eosin (H&E), and observed for histological characteristics.
1.2.5 Observation of Penile Cavernous Fibrosis in Rats
After collecting blood from the abdominal aorta, penile tissue was excised and washed with PBS buffer. The tissue was fully fixed in paraformaldehyde for 72 hours, embedded in paraffin, sectioned, and stained with Masson's trichrome. The relative content of smooth muscle/collagen fibers in the penile cavernous tissue of each group was observed under a microscope.
1.2.6 Measurement of α-SMA Expression Levels in Penile Cavernous Tissue
The penile cavernous tissue was subjected to routine deparaffinization to water, washed with PBS buffer for 5 minutes, and subjected to microwave heat repair in citrate buffer (pH 6.0) for 20 minutes. After cooling to room temperature, the tissue was washed three times with PBS, 3 minutes each time. A 3% hydrogen peroxide solution was applied in a dark room at room temperature for 15 minutes, followed by PBS washing. α-SMA antibody was then added, and the tissue was incubated overnight at 4°C. After returning to room temperature, the tissue was washed with PBS, incubated with a secondary antibody at 37°C for 30 minutes, and washed again with PBS. DAB was applied for color development in the dark, and the reaction was terminated with water. PBS was used as a replacement for the primary antibody as a negative control, and quantitative analysis was conducted with an image analyzer.
1.3 Statistical Methods
The data collected in this study were organized using Microsoft Excel and then analyzed using SPSS 25.0 for statistical testing. The data of each group are expressed as mean ± standard deviation (x̄ ± s). A P-value of less than 0.05 (P < 0.05) was considered statistically significant.
2 Results
2.1 Comparison of Body Weight and Blood Glucose Changes Between the Two Groups
Refer to Table 1.
2.2 Comparison of Penile Erection Rates Between the Two Groups
Refer to Table 2.
2.3 Comparison of Serum Sex Hormone Levels Between the Two Groups
Refer to Table 3.
2.4 Pathological Observation of Testicular Interstitial Tissue Between the Two Groups
The testicular interstitial tissue of the control group appeared normal. The interstitial cells were round or oval in shape, with regular morphology, normal intercellular spacing, orderly arrangement, and no significant reduction in cell number. In contrast, the diabetic group showed testicular interstitial edema, irregular cell morphology, loose intercellular spaces, disordered arrangement, disrupted intracellular structures, and signs of dedifferentiation in some cells, losing structural features. Refer to Figure 1.
| Group | Sample Size (n) | Baseline Body Weight (g) | Body Weight 5 Days After Modeling (g) | Blood Glucose 5 Days After Modeling (mmol/L) | Body Weight Before Sacrifice (g) | Blood Glucose Before Sacrifice (mmol/L) |
|---|---|---|---|---|---|---|
| Diabetes Group | 15 | 220.4 ± 5.5 | 264.2 ± 10.8** | 24.3 ± 5.0** | 316.7 ± 16.4** | 25.2 ± 5.1** |
| Control Group | 5 | 220.0 ± 5.2 | 286.2 ± 10.1 | 4.9 ± 1.3 | 319.1 ± 13.9 | 6.8 ± 1.2 |
Note: Compared to the control group, P < 0.01.
Table 2: Comparison of Penile Erection Rates Between Two Groups
| Group | Sample Size (n) | Positive Cases in Erection Test (n) | Erection Rate (%) |
|---|---|---|---|
| Diabetes Group | 15 | 5 | 33.3** |
| Control Group | 5 | 4 | 80.0 |
Note: Compared to the control group, P < 0.01.
Table 3: Comparison of Serum Sex Hormone Levels Between Two Groups (x̄ ± s)
| Group | Sample Size (n) | T (nmol·L⁻¹) | FSH (IU·L⁻¹) | LH (ng·L⁻¹) |
|---|---|---|---|---|
| Diabetes Group | 15 | 3.38 ± 0.87** | 8.49 ± 3.06** | 489 ± 50.12** |
| Control Group | 5 | 6.04 ± 1.26 | 5.98 ± 1.03 | 386 ± 40.73 |
Note: Compared to the control group, P < 0.01.
2.4 Pathological Observation of Testicular Interstitial Tissue Between Two Groups
In the control group, the testicular interstitial tissue appeared normal, with interstitial cells being round or oval in shape, showing regular morphology, normal intercellular spaces, and orderly arrangement. The number of cells did not show any significant reduction. Compared to the control group, the diabetic group exhibited testicular interstitial edema, irregularly shaped cells, loose intercellular spaces, disordered arrangement, disrupted intracellular structures, and signs of dedifferentiation in some cells, losing their structural features. Refer to Figure 1.
2.5 Observation of Penile Cavernous Fibrosis Between Two Groups
Masson's staining was used to observe the differences in the degree of fibrosis in the penile cavernous tissue between the two groups. In the diabetic group, the proportion of collagen fibers was significantly higher, with an increase in relative content, while the proportion of smooth muscle cells was smaller, with a decrease in relative content. Refer to Figure 2.
Figure 1 HE staining results of testicles in two groups (×10, n = 3)
Figure 2 Masson staining results of corpora cavernosa in two groups (×4, n = 3)
2.6 Expression of α-SMA in corpus cavernosum smooth muscle cells in two groups
Immunohistochemistry was used to detect the expression of α-SMA in the two groups. Compared with the blank group, the brown-yellow stained part in the diabetic group was significantly reduced,that is, the positive cell expression of α-SMA in the diabetic group was significantly reduced. See Figure 3.
Fig. 3 Immunohistochemistry results of corpus cavernosum in two groups (×10, n = 3)
