A time-line profile of the hormone levels during treatment<b>.</b>

<p>For statistical inference, EE was compared with control group, and MP and LD were compared with AR group.</p>&<p>the values (mean±SD) are average of data for 6 animals. Statistical significance is indicated as * P<0.05, ** P<0.005, *** P<0.0005.</p

A time-line profile of sperm count and motility during treatment.

<p>For statistical inference, EE was compared with control group, and MP and LD were compared with AR group.</p>&<p>the values (mean±SD) are average of data for 6 animals. Statistical significance is indicated as * P<0.05, ** P<0.005, *** P<0.0005.</p

Comparison across the treatment groups showed significant increase in ROS upon ethinyl estradiol administration and in the positive control group.

<p>AR group showed slight improvement, but highly significant recovery was seen in the MP and LD groups. Bar diagram shows quantitative assessment of ROS (mean DCF fluorescence). Positive control group was not shown in the bar diagram due to very high ROS value in this group. Data are expressed as Mean ± SD (n = 6). Statistical significance is indicated as **P<0.005, ***P<0.0005 vs. AR.</p

Histological architecture of the testis in different treatment groups.

<p>Control group showed normal testicular histology with the lumen of seminiferous tubules filled with sperm, EE group showed significant compromise in spermatogenesis with almost empty lumens and degeneration of Sertoli cells. AR group showed poor recovery in comparison to complete recovery in MP and LD groups evidenced by densely filled seminiferous tubules and healthy Sertoli cells attached to the basement membrane.</p

Bar diagram comparing the level of apoptosis across the treatment groups.

<p>Ethniyl estradiol administration promoted germ cell apoptosis, resulting in significant increase in dead cells and a corresponding decreased in viable, apoptotic and necrotic cells. Restoration of testicular cells with different fates was seen in the auto-recovery group. MP appeared to be the best in restoring the balance between cell types, but a relatively lesser effect was observed in the LD group. Data are expressed as Mean ± SD (n = 6). Statistical significance is indicated as *P<0.05, **P<0.005, ***P<0.0005 vs. control and ^$$ P<0.005 vs. AR.</p

EE and positive control groups showed significant loss of MMP.

<p>AR group showed recovery of MMP, but a higher level of restoration was seen in the MP and LD groups. The bar diagram depicts the quantitative assessment of MMP across the treatment groups. Data are expressed as Mean ± SD (n = 6). Statistical significance is indicated as ***P<0.0005 vs. control.</p

<em>Mucuna pruriens</em> and Its Major Constituent L-DOPA Recover Spermatogenic Loss by Combating ROS, Loss of Mitochondrial Membrane Potential and Apoptosis

<div><h3>Background</h3><p>The Ayurvedic medicinal system claims <em>Mucuna pruriens</em> (MP) to possess pro-male fertility, aphrodisiac and adaptogenic properties. Some scientific evidence also supports its pro-male fertility properties; however, the mechanism of its action is not yet clear. The present study aimed at demonstrating spermatogenic restorative efficacy of MP and its major constituent L-DOPA (LD), and finding the possible mechanism of action thereof in a rat model.</p> <h3>Methodology/Findings</h3><p>Ethinyl estradiol (EE) was administered at a rate of 3 mg/kg body weight (BW)/day for a period of 14 days to generate a rat model with compromised spermatogenesis. MP and LD were administered in two separate groups of these animals starting 15^th day for a period of 56 days, and the results were compared with an auto-recovery (AR) group. Sperm count and motility, testis histo-architecture, level of reactive oxygen species (ROS), mitochondrial membrane potential (MMP), apoptosis, peripheral hormone levels and testicular germ cell populations were analysed, in all experimental groups. We observed efficient and quick recovery of spermatogenesis in MP and LD groups in comparison to the auto-recovery group. The treatment regulated ROS level, apoptosis, and mitochondrial membrane potential (MMP), recovered the hypothalamic-pituitary-gonadal axis and the number of testicular germ cells, ultimately leading to increased sperm count and motility.</p> <h3>Conclusion/Significance</h3><p><em>M. pruriens</em> efficiently recovers the spermatogenic loss induced due to EE administration. The recovery is mediated by reduction in ROS level, restoration of MMP, regulation of apoptosis and eventual increase in the number of germ cells and regulation of apoptosis. The present study simplified the complexity of mechanism involved and provided meaningful insights into MP/LD mediated correction of spermatogenic impairment caused by estrogens exposure. This is the first study demonstrating that L-DOPA largely accounts for pro-spermatogenic properties of <em>M. pruriens</em>. The manuscript bears CDRI communication number 8374.</p> </div

DNA content analysis of the testicular cell population in the treatment groups.

<p>A significant reduction in spermatid population (1C) after ethinyl estradiol treatment was observed, which was recovered to certain extent in the auto-recovery group. Significant recovery of spermatid population was seen in the MP and LD groups. Data are expressed as Mean ± SD (n = 6). Statistical significance is indicated as *P<0.05 vs. control. SG = Spermatogonia, SSC = Secondary Spermatocytes, PSC = Primary Spermatocytes.</p

Funnel plot.

<p>Plot of precision by log odds ratio using fixed effect model for observed and imputed sets of studies.</p

Flow diagram showing inclusion and exclusion of the studies for meta-analysis.

<p>Studies retrieved upon literature search were subjected to inclusion/exclusion criteria as detailed in the methods section.</p