Effects of Platelet-Rich Plasma on the Oxymetholone-Induced Testicular Toxicity

Abstract

Oxymetholone is one of the anabolic steroids that has widely been used among teenagers and athletes to increase their muscle bulk. It has undesirable effects on male health and fertility. In this study, the therapeutic effects of platelet-rich plasma (PRP) on oxymetholone-induced testicular toxicity were investigated in adult albino rats. During the experiments, 49 adult male albino rats were divided into 4 main groups: Group 0 (donor group) included 10 rats for the donation of PRP, Group I (control group) included 15 rats, Group II included 8 rats that received 10 mg/kg of oxymetholone orally, once daily, for 30 days, and Group III included 16 rats and was subdivided into 2 subgroups (IIIa and IIIb) that received oxymetholone the same as group II and then received PRP once and twice, respectively. Testicular tissues of all examined rats were obtained for processing and histological examination and sperm smears were stained and examined for sperm morphology. Oxymetholone-treated rats revealed wide spaces in between the tubules, vacuolated cytoplasm, and dark pyknotic nuclei of most cells, as well as deposition of homogenous acidophilic material between the tubules. Electron microscopic examination showed vacuolated cytoplasm of most cells, swollen mitochondria, and perinuclear dilatation. Concerning subgroup IIIa (PRP once), there was a partial improvement in the form of decreased vacuolations and regeneration of spermatogenic cells, as well as a reasonable improvement in sperm morphology. Regarding subgroup IIIb (PRP twice), histological sections revealed restoration of the normal testicular structure to a great extent, regeneration of the spermatogenic cells, and most sperms had normal morphology. Thus, it is recommended to use PRP to minimize structural changes in the testis of adult albino rats caused by oxymetholone.

Keywords: albino rat; histology investigation; oxymetholone; platelet-rich plasma; testis; therapeutic effects.

Figure 1

(a) QQ plot of the sample data versus normal standards. (b) Histogram of the rats’ weights at the beginning of the experiment.

Figure 2

A representative diagram of the experimental design.

Figure 3

Photomicrographs of the testis of the control group, showing: (a) Rounded to oval seminiferous tubules (T) lined by spermatogenic cells (Sc) and separated by the interstitial tissue (IT) with blood vessels in between (arrowhead). H&E, ×200. (b) The seminiferous tubule, lined by spermatogenic cells, including spermatogonia (bifid arrow), primary spermatocytes (curved arrow), and spermatids (arrowhead), spermatozoa (Sz). H&E ×400. (c) Spermatogonia (bifid arrow), primary spermatocytes (curved arrow), spermatids (arrowheads), and Sertoli cells (arrow). H&E ×1000. (d) Leydig cells (L) with rounded nuclei, prominent nucleoli (N), and slightly vacuolated acidophilic cytoplasm (C), and myoid cells (thick arrow) and Sertoli cells, with a pyramidal nucleus and prominent nucleolus (arrow). H&E ×1000.

Figure 4

Photomicrographs of a rat testis of group II, showing: (a) Irregular outer boundaries (I), fused tubules (F), widely separated seminiferous tubules (IT), and dilated congested blood vessels (arrowheads). H&E, ×200. (b) Loss of outer boundaries of the tubules (arrows). H&E, ×200. (c) The separation between the basal layer and other spermatogenic cells (*), basal germ cells with very dark nuclei (arrows). H&E, ×400. (d) Deeply inserted spermatozoa in between spermatogenic cells (Sz), desquamated spermatogenic cells in the lumen (Sc), fused outer boundaries of two tubules (F). H&E, ×400. (e) Vacuolated spermatogonia (bifid arrows) and vacuolated primary spermatocytes (curved arrows). H&E, ×1000. (f) Vacuolated interstitial tissue (V) containing Leydig cells (L) with perinuclear vacuolation (N) and highly acidophilic cytoplasm (C). H&E, ×1000.

Figure 5

Photomicrographs of a rat testis of subgroup IIIa, showing: (a) Spermatogonia with dense nuclei and vacuolated cytoplasm (bifid arrows), and partial detachment of the outer boundary of the tubule (arrow). H&E, ×400. (b) Primary spermatocytes with vacuolated cytoplasm (curved arrows), and dark acidophilic cells with fragmented nuclei (arrows). (c) Preserved interstitial cells with some vacuoles (V), Sertoli cells with pyramidal elongated nuclei and prominent nucleoli (arrow), and vacuolated spermatogonia (bifid arrows) (b&C, H&E, ×1000).

Figure 6

Photomicrographs of a rat testis of subgroup IIIb: (a) Spermatozoa in the lumen of the tubules (Sz), and homogenous acidophilic material (*). H&E, ×400. (b) Spermatogonia (bifid arrows), primary spermatocyte (curved arrow), early rounded spermatid (arrowhead), Sertoli cells (arrow). H&E, ×1000.

Figure 7

Photomicrographs of rat testis from different sections showing the PAS reaction: (a–d) basal lamina (arrow), acrosomal cap (arrowhead), and interstitial tissue (*). PAS × 400.

Figure 8

A photomicrograph of rat sperms from group I, showing a head with a characteristic hook (►) and a tail (→) (Eosin, ×1000).

Figure 9

Photomicrographs showing some sperm abnormalities of (group II): (a) a head without a tail (►) and an angulated tail (→), (b) a hookless head (→), (c) a ballooned head (→), (d) a head with a hook at the wrong angle (→), (e) a pin-shaped head (→) with a double tail (►), (f) a tail without a head (►), and a coiled tail (→) (Eosin, ×1000).

Figure 10

Photomicrographs of rat sperms from subgroup IIIa showing a few sperm abnormalities of the single PRP group: (a) an apparently short tail (→) and normal sperm morphology (►), (b) a tail without a head (→), and (c) a separated head fused with a head of another sperm (→) (Eosin, ×1000).

Figure 11

A photomicrograph of rat sperms from subgroup IIIb showing sperms with a nearly normal characteristic head (►) and a long tail (→) (Eosin, ×1000).

Figure 12

Electron micrographs of a rat of the control group, showing: (a) A Sertoli cell resting on basal lamina (BL), nucleus (N) with prominent nucleolus (Nu), mitochondria (M), and electron-dense bodies (E). Mic. Mag. 2500×. (b) Type A spermatogonium, with normal nucleus (N), basal lamina (BL). Mic. Mag. 3000×. (c) Primary spermatocyte, nucleus (N), characteristic vacuolated mitochondria (M). Mic. Mag. 3000×. (d) Spermatid, acrosomal cap (arrow), Golgi apparatus (arrowhead), nucleus (N), mitochondria (M), RER (curved arrows). Mic. Mag. 3000×. (e) Middle pieces of spermatozoa tails showing an axoneme (1), nine outer dense fibers (2), circumferentially arranged mitochondria (3), and a flagellar membrane (4). Mic. Mag. 8000×. (f) Leydig cell with rounded nucleus (N), SER (S), lipid droplets (L), and normal mitochondria (M). Mic. Mag. 3000×.

Figure 13

Electron micrographs of a rat of the oxymetholone group, showing: (a) A Sertoli cell with an area of cytoplasmic loss (*), normal mitochondria (arrows) or partially vacuolated (arrowhead), and secondary lysosomes (zigzag arrows). Mic. Mag. 2000×. (b) Type A spermatogonium, nucleus (N), perinuclear dilatation (arrows), vacuolated cytoplasm (V), myoid cell (My), and dilated SER of neighboring cells (S). Mic. Mag. 3000×. (c) Primary spermatocyte dilated perinuclear spacing (arrows), mitochondria (M), cytoplasmic vacuoles (*). Mic. Mag. 3000×. (d) Spermatid, acrosomal cap (arrows), nucleus (N), mitochondria (M), vacuoles (V), electron-dense bodies (E), spaces between rounded spermatid and adjacent cells (*). Mic. Mag. 2500×. (e) Middle pieces (MP) with excess residual cytoplasm (*). Mic. Mag. 8000×. (f) Leydig cells with dilated perinuclear spaces (arrows), destructed mitochondria (M), and dilated SER (S). Mic. Mag. 3000×.

Figure 14

Electron micrographs of a rat of subgroup IIIa, showing: (a) A Sertoli cell with electron-dense bodies (E) and vacuoles (V). Mic. Mag. 2000×. (b) Type A spermatogonium, myoid cells (My), dilated perinuclear space (arrow), and cytoplasmic vacuole (V). Mic. Mag. 2000×. (c) Primary spermatocyte, nucleus (N), mitochondria (M), electron-dense bodies (E), and vacuoles (V). Mic. Mag. 2000×. (d) Round spermatid, normal acrosomal cap (arrow), Golgi apparatus (arrowhead), nucleus (N), focal discontinuous nuclear membrane (curved arrow), normal mitochondria (M), swollen mitochondria (m), and dilated RER (R). Mic. Mag. 3000×. (e) Middle pieces (MP) and principal pieces (PP) with excess residual cytoplasm (*). Mic. Mag. 6000×.

Figure 15

Electron micrographs of a rat of subgroup IIIb, showing: (a) A Sertoli cell with nucleus (N), mitochondria (M), and basal lamina (BL). Mic. Mag. 3000×. (b) Type A spermatogonium, nucleus with areas of heterochromatin (arrowhead) and focal perinuclear dilatation (arrows), and myoid cell (My). Mic. Mag. 2000×. (c) Primary spermatocyte, nucleus (N), mitochondria (M), and nuclear membrane shows localized dilatation (arrow). Mic. Mag. 2500×. (d) Round spermatid, normal nucleus (N), intact mitochondria (M), and dilated RER (arrows). Mic. Mag. 3000×. (e) Intact middle pieces (MP), principal pieces (PP), and end pieces (EP). Mic. Mag. 5000×.