|Year : 2021 | Volume
| Issue : 3 | Page : 172-181
Role of maternal nutritional supplementation on the hormonal profile and immunohistochemical analysis of testicular development of fetal rats
Taiwo O Kusemiju1, Olasunmbo O Afolayan1, Babatunde Ogunlade2
1 Department of Anatomy, College of Medicine, University of Lagos, Lagos, Nigeria
2 Department of Anatomy, College of Medicine, University of Lagos, Lagos; Department of Human Anatomy, Behavioral and Aging Unit, Federal University of Technology, Akure, Ondo, Nigeria
|Date of Submission||24-Jun-2021|
|Date of Decision||26-Jul-2021|
|Date of Acceptance||31-Jul-2021|
|Date of Web Publication||30-Nov-2021|
Dr. Olasunmbo O Afolayan
Department of Anatomy, College of Medicine, University of Lagos, Lagos
Source of Support: None, Conflict of Interest: None
Introduction: This study elucidated the role of maternal microelement supplementation on fetal testicular development. Materials and Methods: Twenty-eight Wistar rats (21 females and 7 males) were randomly divided into seven groups comprising three females to one male: Group A – standard feed and water; Group B – diet deficient in all the micronutrients (Fe, Cu, Se) and water; Group C – diet fortified with 0.2 mg/kg of selenium and water; Group D – diet enriched with 40 mg/kg of iron and water; Group E – diet fortified with 8 mg/kg of copper and water; and Group F – diet enriched with all nutrients and water. All administration was via oral gavage; thereafter, animals were sacrificed at day 20 of pregnancy. Placenta measurements, testes, and blood serum were obtained for analysis. Results: The results showed statically a significant decrease (p<0.05) in placental and fetal weight (WPF), the distance between fetus and mother (DFM), the concentration of trace elements; FSH and LH levels among chaff only group when compared to the Control group. Testicular histomorphology and immunohistochemical studies of the animals in the chaff alone diet showed mild fetal Leydig cells in the interstitium, primitive germ cells in the testicular cord, pre-Sertoli cell necrosis, and decreased positive expression compared with the control. Conclusion: Administration of single and combined doses of nutritional supplements diet significantly preserved the fetal parameters, hormone profile, and histochemical analysis of the testis.
Keywords: Fetal outcome, hormone profile, immunohistochemistry, nutritional trace elements, testis
|How to cite this article:|
Kusemiju TO, Afolayan OO, Ogunlade B. Role of maternal nutritional supplementation on the hormonal profile and immunohistochemical analysis of testicular development of fetal rats. Niger J Exp Clin Biosci 2021;9:172-81
|How to cite this URL:|
Kusemiju TO, Afolayan OO, Ogunlade B. Role of maternal nutritional supplementation on the hormonal profile and immunohistochemical analysis of testicular development of fetal rats. Niger J Exp Clin Biosci [serial online] 2021 [cited 2022 Jun 26];9:172-81. Available from: https://www.njecbonline.org/text.asp?2021/9/3/172/331555
| Introduction|| |
Nutrition is regarded as the intake of food necessary for growth, function, and health. A diet that provides all essential nutrients in optimal amounts and proportion is classified as proper nutrition. Poor nutrition is regarded as a diet that lacks nutrients either from imbalance or overall insufficient food intake. Appropriate fetal growth and development of a developing fetus are supported by adequate nutritional status. Dietary energy and nutrients requirement is said to be increased during pregnancy to support increased maternal metabolism, blood volume, and delivery of nutrients to the fetus. Exposure to poor nutrition (either over- or undernutrition) during development can affect the programming of adult anatomy, physiology, and metabolism. The structure and function of testes, prostate glands, and gonads are said to be affected by maternal malnutrition leading to impairment of reproductive capacity of the male offspring. More so, maternal nutrition and lifestyle have been said to play a significant role in the proper function of the reproductive system, with poor maternal nutrition being the leading cause of infertility affecting 60–80 million couples worldwide., Studies have also reported that male factor infertility contributes to more than 40% of infertility cases globally. Evidence has it that decreased sperm count has been linked to the crucial role of nutrition.,
Animals and human studies have shown that male obesity, high-fat diet, maternal nutrition, and impaired productivity affect the molecular and physical structure of sperm, as well as the health of the developing fetus and subsequent offspring. Rato et al. thereby affirmed that high-energy diets affect testicular metabolism, and testicular physiology is sensitive to alteration of whole-body metabolism. Decreased sperm count has been linked to the disruption of testicular metabolism.
Deficiency in vitamins or minerals affects about two billion people globally, which include children and women of reproductive age; pregnant women are also vulnerable to such deficiencies. Micronutrients such as selenium are said to play a crucial role in spermatogenesis to protect sperm against oxidative substances. Deficiency in selenium in recent studies has been linked with smaller testicles, and in long-term sperm impairment through spermatogenesis and sperm maturation process. Shi et al. confirmed that selenium supplementation promotes testicular and semen glutathione peroxidase activity, as well as protects the membrane system integrity and proliferation of spermatogonial stem cells.
Similarly, copper has been reported to play a significant role in spermatogenesis and male infertility in Wistar rats. Copper is also involved in spermatozoa motility and may also act at the pituitary receptors which control the release of luteinizing hormone (LH). A significant decrease in spermatozoa concentration, motility, and viability indicates the adverse effect of copper in male fertility.
Testicular abnormalities such as testicular atrophy, morphological changes in the testis, impaired spermatogenesis, epididymal lesions, and impaired reproductive function have been linked to deficiency of iron. Iron deficiency is reported to be connected with beta-thalassemia which is a leading cause of male infertility, sexual dysfunction, delay in pubertal growth, and inadequate sexual development. Biological markers for testicular development such as parvalbumin-a calcium-binding marker have been used to assay the importance of calcium during testicular development. Calcium has been reported to play a significant role in sperm motility, acrosome reaction, and capacitation. On the other hand, transferrin, a biological marker, takes part in iron homeostasis as a protein carrying iron to the target cell. This study is aimed at investigating the role of maternal nutritional diet supplementation on testicular development.
| Materials and Methods|| |
Twenty-eight Wistar rats (21 females and 7 males) weighing 150 g–200 g were obtained from the breeding stock of Jumorak farm at Iwo, Osun state. The rats were collected in the isolated cages and allowed to acclimatize for 14 days in the experimental house of the Department of Anatomy, College of Medicine of the University of Lagos, before the commencement of the experiment. The rats were maintained under the standard natural photoperiodic condition of 12 h of light alternating with 12 h of darkness (i.e., L:D; 12 h: 12 h photoperiod) at room temperature (27°C–30°C) and humidity of 65 ± 5. They were allowed unrestricted access to water and rat chow. Picric acid was used to tag the animals as head (H), body (B), and tail (T) for possible identification. The processes of protocols using the experimental animals were in accordance with the Guide for the Care and Use of Laboratory Animals and approved by the Health Research Ethics Committee of the College of Medicine, University of Lagos.
The rats were randomly divided into seven groups comprising three females to one male, labeled as groups A–F. Vaginal smear was carried out to confirm their estrus cycle before and after mating. Vaginal plug was verified for possible pregnancy and was recorded as gestational day zero (GD 0) before starting the treatment. Group A received standard feed and water; Group B was fed with a diet deficient in all the micronutrients (Fe, Cu, Se) and water; Group C was fed with 0.2 mg/kg of selenium and water; Group D was fed with 40 mg of iron per kg and water; Group E was fed with 8 mg of copper per kg and water; and Group F was fed with a diet fortified with all nutrients (Fe, Cu, Se) and water.
Micronutrient supplements were administered to the appropriate group at gestational day 5 (GD 5) in utero, targeting the critical period of testicular development which was recorded to be embryonic day 15.5–18.5 (e15.5–e18.5). All animals were observed for any anomalies, illness, and physical anomalies. The weights of the animals were recorded weekly and were assed at prenatal day 19.
Quantification of metals in the rat diet using an atomic absorbent spectrophotometer is given in [Table 1].
|Table 1: Showing the mean and standard error of mean values of the weight of the placenta (WP), weight of placenta and fetus (WP&F), distance between placenta and fetus (DP&F), and distance between fetus and mother (DF&M) of the different animal groups|
Click here to view
Testicular histology preparation
Rats were harvested and fixed in Bouin's fluid for 24 h before being transferred into 70% alcohol for dehydration. The tissues passed through 90% and absolute alcohol and xylene for different durations before being transferred into molten paraffin wax for 1 h each in an oven at 65°C for infiltration. The tissues were embedded and serial sections cut on a rotary microtome set at 5 μ were performed. Each testis was picked up with slides and allowed to dry on hot plates for 2 min. The slides were dewaxed with xylene and passed through absolute alcohol (two changes), 70% alcohol, 50% alcohol, (in that order), and then in water for 5 min. The slides were then stained with hematoxylin and eosin, mounted in DPX, and photomicrographs were taken at a magnification of ×400 on a Leica DM750 microscope.
Quantification of metal in the testis
The iron, zinc, copper, and selenium contents in the tissues were determined after digestion in nitric acid (HNO3) in the microwave digestion system. After that, the concentrations of iron, zinc, copper, and selenium in the mineral solutions were measured using the flame atomic absorption spectrometry method.
The levels of LH and follicle-stimulating hormone (FSH) were measured using available immunoassay (ELISA) kits (Randox Laboratories, United Kingdom) according to manufacturer instructions.
Immunohistochemical analysis of transferrin and parvalbumin in the testis
Tissue was deparaffinized and rehydrated, immersed in xylene for 10 min, excess liquid was removed, and then dipped in anhydrous ethanol for 3 min. The slides were immersed in 85% ethanol for 3 min and rinsed in de-ionized water, and then rinsed in PBS buffer for 3 min. After deparaffinization and rehydration, antigen retrieval was done, 1× ethylenediaminetetraacetic acid antigen repair was added to tissues after which slides were removed and rinsed with PBS for 3 min (Protocol for immunohistochemistry kit).
The data were presented as mean ± standard error of mean and were analyzed using one-way analysis of variance followed by a post hoc Tukey's test, which was performed using GraphPad Prism version 8.00 for Windows (GraphPad Software, San Diego, USA). A result of P < 0.05 was considered statistically significant.
| Results|| |
Effects of microelement enriched nutritional dietary on fetal measurements in rats
The results revealed that rats that consumed chaff showed a significant decrease (P < 0.05) in placenta weight and fetal weight (WPF) as well as the distance between fetus and dams (DFM) compared to the control group [Table 1]. The rats that were fed with Se, Fe, and Cu in their diets independently and the group that consumed a diet supplemented with the combination of Se, Fe, and Cu showed a significant increase (P < 0.05) in the placenta and fetal weight (WPF), the distance between placenta and fetus (DPF), and distance between fetus and mother (DFM) when compared with the chaff diet only group [Table 1], although there was a significant decrease (P < 0.05) in weight of the placenta and the fetus (WPF), the distance between placenta and fetus (DPF), and distance between fetus and mother (DFM) among the rats administered with a diet containing independent dose of Se, Fe, and Cu when compared with a diet supplemented with Se, Fe, and Cu [Table 1].
Concentration of microelements in the testis of rats exposed to supplemented diets
The results revealed that rats treated with chaff only showed a significant decline (P < 0.05) in Se, Fe, and Cu concentration relative to the control group [Table 2]. However, the rats that consumed a diet containing Se, Fe, and Cu and a diet containing a mixed combination of the microelements showed a significant increase (P < 0.05) in Se, Fe, and Cu concentration when compared with the chaff diet only group [Table 3], although there was a significant decrease (P < 0.05) in Se, Fe, and Cu concentration among the rats that fed on diet containing Se, Fe, and Cu independently when compared with a mixed combination of the microelements in their diet [Table 2].
|Table 2: Showing the mean and standard error of mean of copper concentration, iron concentration, and selenium concentration in each animal groups.(ppm)|
Click here to view
|Table 3: Showing Follicle stimulating hormone (FSH) and Luteinizing hormone (LH)|
Click here to view
Effects of microelement supplementation on follicle-stimulating hormone and luteinizing hormone in male rats
The results revealed that rats treated with chaff only showed a significant decline (P < 0.05) in FSH and LH levels relative to the control group [Table 3]. However, the rats that consumed Se, Fe, and Cu independently in their diets showed a significant increase (P < 0.05) in FSH and LH levels when compared with the chaff diet only group [Table 3], although there was a significant decrease (P < 0.05) in FSH and LH levels among the rats that were fed with a mixed combination of the microelements when compared with single doses of nutritional supplements diet (Se, Fe, and Cu) [Table 3].
Effects of microelement supplementation on the histomorphology of testis in rats
The photomicrograph of the testicular histomorphology of the animals in the chaff diet (Group B) [Figure 1] showed mild fetal Leydig cells in the interstitium, primitive germ cells in the testicular cord, and pre-Sertoli cell necrosis when compared with the control (Group A). However, in [Figure 1], the testis of the control had similar characteristics with the single doses of nutritional supplements diet (Se, Fe, and Cu) and mixed combination of the microelements with diet showing testicular cords, fetal Leydig cell in the interstitium, basal lamina, darkly stained primitive germ cell (gonocyte), and pre- Sertoli cells More Details. The testicular section of the group that received a mixed combination of the supplements with diet showed more restored cyto-architecture of the testicular morphology compared to the single doses of nutritional supplements diet (Se, Fe, and Cu) [Figure 2] and [Figure 3], [Figure 4], [Figure 5], [Figure 6].
|Figure 1: B1, B2 photomicrograph of embryonic day 19 of fetal testis in chaff group following in utero exposure to deficient diet showing mild fetal leydig cells in the interstitium (IS), uneven shaped testicular cords (TC), basal lamina (arrow head), primitive germ cell in testicular cord (black arrow), and pre-sertoli cell(red arrow). Magnification of x100(top); x400(bottom) under light microscope.Stained with hematoxylin and eosin (H&E).|
Click here to view
|Figure 2: A3, A4 photomicrograph of embryonic day 19 fetal testes in control showing testicular cords(TC), fetal leydig cell in the interstitium (IS), basal lamina(arrow head), darkly stained primitive germ cell (gonocyte) (black arrow) and pre-sertoli cells(red arrow). Magnification x100(top) and x400(bottom) under light microscope. Stained with hematoxylin and eosin (H&E).|
Click here to view
|Figure 3: C1, C2 photomicrograph of embryonic day 19 of a fetal testis in copper diet group, following inutero exposure to copper diet showing testicular cord(TC), mild fetal leydig cells in the interstitium compartment, basal lamina (arrow head), reduced primitive germ cell in the cord (black arrow), and pre-sertoli cell(red arrow). Magnification x100(above); x400(below).Under light microscope. Stained with hematoxylin and eosin (H&E)|
Click here to view
|Figure 4: D1, D2 photomicrograph of embryonic day 19 fetal rat testis in selenium or inutero exposure to selenium diet showing testicular cord(TC), well defined basal lamina(arrow head) and increased number of fetal leydig cells in the interstitium compartment, more primitive germ cell(black arrow), and sertoli cell (red arrow). Magnification x100 (above); x400(below).Under light microscope.Stained with hematoxylin and eosin (H&E)|
Click here to view
|Figure 5: E1, E2 photomicrograph of embryonic day 19 fetal rat testis in iron group or inutero exposure to iron diet showing testicular cord(TC), increase fetal leydig cell in the interstitial compartment (IS), more primitive germ cell in the cord (black arrow), pre-sertoli cell (red arrow), and basal lamina (arrow head). Magnification x100 (above); x400(below) under light microscope.Stained with hematoxylin and eosin (H&E)|
Click here to view
|Figure 6: F1, F2 photomicrograph of embryonic day 19 fetal rat testis in mix diet group showing the testicular cord (TC), increased leydig cell in the interstitial compartment (IS), increased primitive germ cells in the cord(black arrow) well defined basal lamina (arrow head), and and pre-sertoli cells (red arrow). Magnification x100(above); x400(below). Under light microscope stained with hematoxylin and eosin (H&E)|
Click here to view
Effects of nutritional diet supplement on testicular immunohistochemical analysis (parvalbumin) in normal and experimental rats
The photomicrograph of the testicular immunohistochemistry (parvalbumin) of the animals in the chaff only diet (Group B) [Figure 7] showed mild localization in the interstitial compartment of fetal Leydig cells and pre-Sertoli cells, and primitive germ cells when compared with the control (Group A) [Figure 8]. However, testicular photomicrograph of the control section had similar characteristics with the single doses of nutritional supplement diet (Se, Fe, and Cu) and mixed combination of the supplements with diet showing sparse localization in the interstitial compartment of fetal Leydig cells and pre-Sertoli cells, and the primitive germ cell [Figure 8] and [Figure 9], [Figure 10], [Figure 11], [Figure 12].
|Figure 7: B1, B2. Parvalbumin localization by immunohistochemistry in fetal rat testis on embryonic day 19 from chaff group fetus showing the region of mild localization in the interstitial compartment of fetal leydig cell (circle area) and pre-sertoli cells (rectangle area), and arrow pointing to primitive germ cell|
Click here to view
|Figure 8: A1, A2. Parvalbumin localization by immunohistochemistry in the fetal rat testis on embryonic day 19 from control fetus the interstitial compartment of the testis contained sparse number of stained fetal leydig cell (circle area), and pre-sertoli cell (rectangle area), and arrow showing primitive germ cell|
Click here to view
|Figure 9: C1, C2. Parvalbumin localization by immunohistochemistry of embryonic day 19 fetal rat testis from copper diet group fetus showing sparse localization in the interstitial compartment of fetal leydig cell (circle area) and pre-sertoli cell (rectangle area), with the arrow pointing to primitive germ cell|
Click here to view
|Figure 10: D1, D2. Parvalbumin localization by immunohistochemistry of embryonic day 19 fetal rat testis selenium diet group showing positive expression in interstitial compartment of fetal leydig cells(circle region) and pre-sertoli cells (rectangle region) with the arrow pointing to primitive germ cell|
Click here to view
|Figure 11: E1, E2. Parvalbumin localization by immunohistochemistry of embryonic day 19 fetal rat testis from iron diet fetus group showing sparse localization in the interstitial compartment of fetal leydig cell (rectangle region) and pre-sertoli cell (circle region), with the arrow pointing toward the primitive germ cell|
Click here to view
|Figure 12: F1, F2. Parvalbumin localization by immunohistochemistry of embryonic day 19 fetal rat testis from mix diet group fetus showing sparse localization in the interstitial compartment of fetal leydig cells (circle region) and pre-sertoli cells (rectangle region) with the arrow pointing towards the primitive germ cell|
Click here to view
Effects of nutritional diet supplement on testicular immunohistochemical analysis (transferrin) in normal and experimental rats
The photomicrograph of the testicular immunohistochemistry (transferrin) of the animals in the chaff only diet (Group B) [Figure 13] showed more positive expression in the preSertoli cell and primitive germ cell when compared with the control (Group A) [Figure 14]. However, testicular photomicrograph of the control section had similar characteristics with the single doses of nutritional supplements diet (Se, Fe, and Cu) and mixed combination of the supplements with diet showed positive expression in preSertoli cell and increased number of germ cell [Figure 14] and [Figure 15], [Figure 16], [Figure 17], [Figure 18].
|Figure 13: B1, B2. Localization of transferrin by immunohistochemistry of embryonic day 19 fetal rat testis from the chaff fetus showing more positive expression in the pre-sertoli cell and primitive germ cell (Rectangle)|
Click here to view
|Figure 14: A1, A2; Transferrin localization by immunohistochemistry of embryonic day 19 fetal rat testis from the control fetus showing positive localization in the pre-sertoli cell (rectangle) and the arrow pointing to primitive germ cell|
Click here to view
|Figure 15: C1, C2. Localization of transferrin by immunohistochemistry of embryonic day 19 fetal rat testis from copper diet group fetus testis showing more positive expression in the pre-sertoli cell (rectangle) and arrow pointing to primitive germ cell. Reduced number of primitive germ cell evidence|
Click here to view
|Figure 16: D1, D2. Localization of transferrin by immunohistochemistry of embryonic day 19 of fetal rat testis from the selenium diet group fetus testis showing positive localization in the pre-sertoli cells and more prominent primitive germ cell (arrow pointing to primitive germ cell)|
Click here to view
|Figure 17: E1, E2. Localization of transferrin by immunohistochemistry of embryonic day 19 from iron diet group fetus testis showing positive expression in pre-sertoli cell (rectangle) and increased number of germ cell evidence.Arrow pointing to primitive germ cell|
Click here to view
|Figure 18: F1, F2 localisation of transferrin by immunohistochemistry of embroyonic day 19 fetal rat testis from mix diet group showing positive expression of transferrin in pre-sertoli cells(rectangle) and arrow pointing to primitive germ cell|
Click here to view
| Discussion|| |
Pregnancy accounts for an increased maternal volume and an increased need for nutrients for the development and growth of the fetus and placenta. Even though a variety of factors can be said to contribute to the chances of developmental abnormalities, there is a high event that human pregnancies can be an outcome essentially impaired by less optimal maternal nutritional status, resulting in poor nutrition being the leading cause of preventable birth defects. Depleted micronutrient concentration may arise as a result of low food intake, low nutrient-dense food choice, poor eating habit, and food intolerance.
It has been established that during pregnancy, the concentration of selenium in the blood decreases significantly. Changes in selenium homeostasis during pregnancy were affirmed to be as a result of increased demand for oxygen in the body of the mother and developing fetus. Selenium has been reported to be transported passively in the maternal-fetal direction, suggesting that the placenta could act as a regulator of fetal selenium transfer. In this present study, placenta selenium value was significantly higher in the fetuses exposed to selenium diet compared to other groups. Evidence from [Table 1] shows that there was no statistical significance recorded between control and other experimental groups, but there was statistical significance between the mixed diet group and the copper, iron, and selenium diet group. Copper has been established to act as a co-factor in antioxidant enzymatic activities, which shields the fetus from any undue impairment. Therefore, the deficiency of this metal in the placenta acts as one of the detrimental factors responsible for the depletion of antioxidant activities, which may result in oxidative stress in trophoblastic placenta tissue and is a curse for fetus growth, responsible for early pregnancy and low birth weight. On the other hand, iron deficiency during pregnancy has been reported to be a major problem in developing countries and is strongly associated with low birth weight.
The weight of the placenta and fetus showed a statistically significant increase between chaff and other experimental groups. When comparing the weight of the fetus and placenta between the iron diet group and the control and mixed diet groups, it was statistically increased. The copper diet group with control and mixed diet groups showed a statistically significant increase in the weight of the fetus and placenta. Placenta has been reported to be essential for the establishment of fetal development in utero and for the establishment of adequate birth iron stores to sustain growth in early infancy, and it aids in the optimal transfer of iron between the mother and the fetus. In contrast, Sferruzzi-Perri and Camm deduced that fetal and placenta weight that are on average lower than the ad-libitum controls have been linked to reducing maternal food intake by 15%–50% for the majority of pregnancy in rats and mice.
The distance between placenta and fetus showed a very highly statistical significant between iron and control, copper diet, selenium diet, and mixed diet when compared with chaff. This result suggested that maternal iron diet was transported via the placenta resulting in a considerable increase in the distance between the fetus and the placenta. Iron deficiency in pregnancy leading to anemia is caused by the ability of the fetus to obtain iron requirement in one-way direction even from the iron deficient mothers.
The distance between placenta and mother showed a very high significance when comparing chaff with other diets. The results of the distance between the placenta and mother showed a significant increase in the control group, selenium fortified diet group, and mixed diet group suggesting the role this diet played in development. No literature has recorded the significance of this diet and any reproductive significance associated with the distance between the placenta and mother.
Furthermore, the metal analysis result showed that the copper concentration in the testis was increased in the copper diet group compared to other experimental groups and the chaff group also showed an increase in the copper concentration deducing that maternal copper diet increases the concentration of copper in the testis of the fetus exposed to copper diet and chaff (a deficient diet). The increased copper level in the testis suggested that the pregnant mother was exposed to the right quantity of copper. Pal also confirmed that male rats fed low copper diet exhibited decreased copper levels in the testis. In addition, Zhang et al. also confirmed that sublethal exposure to copper induces sperm head malformation, and influences both mRNA and protein levels of glutathione peroxidase 5 and androgen receptor that are related to copper. In addition, iron deficiency has been reported to have important health complications for the growing fetus, besides increasing the risk of perinatal mortality. The pregnant dam in this study was exposed to the right quantity of iron diet suggesting its increase in the testis. There was an increased concentration of selenium in the selenium diet group compared to control and other experimental groups, this result indicated the role selenium plays in testicular development. According to Eroglu et al., selenium supplementation caused a significant increase in testicular development.
In agreement with this result, it was affirmed that the animals that were subjected to selenium supplementation showed an increase in LH concentration. FSH analysis showed a significant increase between mixed diet group and iron diet group suggesting the importance of maternal iron diet and a mixed diet (standard diet) in the reproductive function of the testis. In contrast to our result, iron overload in hypogonadism patients showed a decrease in the gonadotrophin hormone (FSH and LH).
Histological analysis of the fetal testis of normal, single, and mixed doses of trace elements diet showed an increased number of fetal Leydig cells in the interstitium, well-defined testicular cords and basal lamina, and an increased number of primitive germ cells when compared to chaff group. Di Bona et al. confirmed that animals exposed to high doses of iron (100 mg/kg) showed a decrease in the germinal epithelium resulting in loss in germ cells. The finding of Azman and Senol also deduced that animals exposed to iron showed breakage in the seminiferous tubule epithelium, decrease in epithelial thickness, and decrease in germ cells, severe edema, and congestion in the interstitial area. All these were in contrast to our findings in the histology of the fetus exposed to iron diet.
In contrast to the result gotten from the histology of fetal testis exposed to selenium diet, Bano et al. deduced that decreased number of Leydig cells and poor testicular development were associated with animals exposed to selenium diet. The copper diet group showed a mild fetal Leydig cell and reduced primitive germ cells. The result obtained from the histology of the fetal testis exposed to copper diet was in agreement with the results of Chen et al. that affirmed testis of animals exposed to higher amount of copper showed a decreased number of spermatogonia. This result suggested the role maternal iron, copper, selenium, and standard (mixed diet) play in the development of testicular development.
Immnuno-expression of transferrin showed a positive expression in the testis of the fetuses exposed to copper diet, selenium diet, iron diet, mixed diet, and chaff when compared to the control group, thereby implying that transferrin secreted by the Sertoli cells was able to transport iron to the germinal epithelium and the primitive gonad (gonocytes). The result is in affirmation with Gunes et al. that deduced that the transferrin receptor is present in the region of the Sertoli cells. Furthermore, immune expression of parvalbumin was positively expressed in fetus exposed to selenium diet compared to control and other experimental groups, this shows that maternal selenium diet supported the expression of this testicular marker and that the intracellular calcium was able to bind with parvalbumin which, in turn, transports calcium to the developing primitive germ cells (Gonocytes). Tvrdá et al. affirmed that parvalbumin was recently found to be expressed in steroidogenic cells such as adrenal glands and Leydig.
| Conclusion|| |
This study assessed the effect of maternal nutrition on the testicular development of a Wistar rat. The evidences from this study deduced that the metal concentration of trace elements (selenium, copper, and iron) in the testis of the fetus was higher, showing the importance of these elements in the testicular development. Therefore, maternal nutrition supplementation of selected trace elements has successfully played a significant role in preserving the cyto-architecture and functional integrity of the testis in the development of the fetuses.
Financial support and sponsorship
Conflicts of interest
There are no conflicts of interest.
| References|| |
Hujoel PP, Lingström P. Nutrition, dental caries and periodontal disease: A narrative review. J Clin Periodontol 2017;44 Suppl 18:S79-84.
Hanson MA, Bardsley A, De-Regil LM, Moore SE, Oken E, Poston L, et al.
The International Federation of Gynecology and Obstetrics (FIGO) recommendations on adolescent, preconception, and maternal nutrition: “Think Nutrition First”. Int J Gynaecol Obstet 2015;131 Suppl 4:S213-53.
Branca F, Piwoz E, Schultink W, Sullivan LM. Nutrition and health in women, children, and adolescent girls. BMJ 2015;351:h4173.
Dunlap KA, Brown JD, Keith AB, Satterfield MC. Factors controlling nutrient availability to the developing fetus in ruminants. J Anim Sci Biotechnol 2015;6:16.
Brett KE, Ferraro ZM, Yockell-Lelievre J, Gruslin A, Adamo KB. Maternal-fetal nutrient transport in pregnancy pathologies: The role of the placenta. Int J Mol Sci 2014;15:16153-85.
Rinaldi JC, Santos SA, Colombelli KT, Birch L, Prins GS, Justulin LA, et al.
Maternal protein malnutrition: Effects on prostate development and adult disease. J Dev Orig Health Dis 2018;9:361-72.
Kumar N, Singh AK. Trends of male factor infertility, an important cause of infertility: A review of literature. J Hum Reprod Sci 2015;8:191-6.
] [Full text]
Fleming TP, Eckert JJ, Denisenko O. The role of maternal nutrition during the periconceptional period and its effect on offspring phenotype. Adv Exp Med Biol 2017;1014:87-105.
Agarwal A, Mulgund A, Hamada A, Chyatte MR. A unique view on male infertility around the globe. Reprod Biol Endocrinol 2015;13:37.
Ilacqua A, Izzo G, Emerenziani GP, Baldari C, Aversa A. Lifestyle and fertility: The influence of stress and quality of life on male fertility. Reprod Biol Endocrinol 2018;16:115.
Rato L, Alves MG, Dias TR, Lopes G, Cavaco JE, Socorro S, et al
. High-energy diets may induce a pre-diabetic state altering testicular glycolytic metabolic profile and male reproductive parameters. Andrology 2013;1:495-504.
Wennink JM, Delemarre-van de Waal HA, Schoemaker R, Schoemaker H, Schoemaker J. Luteinizing hormone and follicle stimulating hormone secretion patterns in girls throughout puberty measured using highly sensitive immunoradiometric assays. Clin Endocrinol (Oxf) 1990;33:333-44.
Darnton-Hill I, Mkparu UC. Micronutrients in pregnancy in low- and middle-income countries. Nutrients 2015;7:1744-68.
Walczak-Jedrzejowska R, Wolski JK, Slowikowska-Hilczer J. The role of oxidative stress and antioxidants in male fertility. Cent European J Urol 2013;66:60-7.
Monfared YK, Khodabandehloo E. Effects of selenium on various sperm parameters in varicocele rats. World family medicine/middle east journal of family medicine 2018;16(2)270-4.
Shi L, Zhao H, Ren Y, Yao X, Song R, Yue W. Effects of different levels of dietary selenium on the proliferation of spermatogonial stem cells and antioxidant status in testis of roosters. Anim Reprod Sci 2014;149:266-72.
Liu JY, Yang X, Sun XD, Zhuang CC, Xu FB, Li YF. Suppressive effects of copper sulfate accumulation on the spermatogenesis of rats. Biol Trace Elem Res 2016;174:356-61.
Roychoudhury S, Nath S, Massanyi P, Stawarz R, Kacaniova M, Kolesarova A. Copper-induced changes in reproductive functions: In vivo
and in vitro
effects. Physiol Res 2016;65:11-22.
Sakhaee E, Abshenas J, Emadi L, Azari O, Kheirandish R, Samaneh A. Effects of vitamin C on epididymal sperm quality following experimentally induced copper poisoning in mice. Comp Clin Pathol 2014;23:181-6.
Bomhard EM, Gelbke HP. Hypoxaemia affects male reproduction: A casestudy of how to differentiate between primary and secondary hypoxic testicular toxicity due to chemical exposure. Arch Toxicol 2013;87:1201-8.
Lanzkowsky P. Iron-deficiency anemia. In: Lanzkowsky's Manual of Pediatric Hematology and Oncology. Europe PMC; 2016: 21(2) 69 83.
KosakaT, Yasuda S, Kosaka K. Calcium-binding protein, secretagogin, characterise novel groups of interneurons in the rat striatum. Neurosci Res 2017;119:53-60.
Ayaz O, Howlett SE. Testosterone modulates cardiac contraction and calcium homeostasis: Cellular and molecular mechanisms. Biol Sex Differ 2015;6:9.
Wallace DF. The regulation of iron absorption and homeostasis. Clin Biochem Rev 2016;37:51-62.
Sarkar M. Evaluation of the anti-inflammatory activity of methanolic leaf extract of Mangifera indica L. (Anacardiaaceae) in rats. Int J Drug Dev Res 2015;7:26-30.
Kermani ES, Nazari Z, Mehdizadeh M, Shahbazi M, Golalipour MJ. Gestational diabetes influences the expression of hypertrophic genes in left ventricle of rat's offspring. Iran J Basic Med Sci 2018;21:525-8.
Lara NL, van den Driesche S, Macpherson S, França LR, Sharpe RM. Dibutyl phthalate induced testicular dysgenesis originates after seminiferous cord formation in rats. Sci Rep 2017;7:2521.
Liao Z, Cao H, Dai X, Xing C, Xu X, Nie G, et al.
Molybdenum and Cadmium exposure influences the concentration of trace elements in the digestive organs of Shaoxing duck (Anas platyrhyncha
). Ecotoxicol Environ Saf 2018;164:75-83.
Gernand AD, Schulze KJ, Stewart CP, West KP Jr., Christian P. Micronutrient deficiencies in pregnancy worldwide: Health effects and prevention. Nat Rev Endocrinol 2016;12:274-89.
Mennitti LV, Oliveira JL, Morais CA, Estadella D, Oyama LM, Oller do Nascimento CM, et al.
Type of fatty acids in maternal diets during pregnancy and/or lactation and metabolic consequences of the offspring. J Nutr Biochem 2015;26:99-111.
Troesch B, Biesalski HK, Bos R, Buskens E, Calder PC, Saris WH, et al.
Increased intake of foods with high nutrient density can help to break the intergenerational cycle of malnutrition and obesity. Nutrients 2015;7:6016-37.
Mao J, Vanderlelie JJ, Perkins AV, Redman CW, Ahmadi KR, Rayman MP. Genetic polymorphisms that affect selenium status and response to selenium supplementation in United Kingdom pregnant women. Am J Clin Nutr 2016;103:100-6.
Kosik-Bogacka D, Łanocha-Arendarczyk N, Kot K, Malinowski W, Szymański S, Sipak-Szmigiel O, et al.
Concentrations of mercury (Hg) and selenium (Se) in afterbirth and their relations with various factors. Environ Geochem Health 2018;40:1683-95.
Sakamoto M, Chan HM, Domingo JL, Koriyama C, Murata K. Placental transfer and levels of mercury, selenium, vitamin E, and docosahexaenoic acid in maternal and umbilical cord blood. Environ Int 2018;111:309-15.
Hefnawy AE, El-Khaiat H. Copper and animal health (importance, maternal fetal, immunity and DNA relationship, deficiency and toxicity). Int J Agro Vet Med Sci 2015;9:195-211.
Thaker R, Hinda O, Idrish S, Sunel K. Correlation of copper and zinc in spontaneous Abortion. Int J Fertility 2019;13:97-101.
Singh R, Chauhan R, Nandan D, Singh H, Gupata SC, Bhatnagar M. Morbidity profile of women during pregnancy: A hospital record-based study in Western UP. IJCH 2012;24:342-6.
Cao C, Fleming MD. The placenta: The forgotten essential organ of iron transport. Nutr Rev 2016;74:421-31.
Sferruzzi-Perri AN, Camm EJ. The programming power of the placenta. Front Physiol 2016;7:33.
Orlandini C, Torricelli M, Spirito N, Alaimo L, Di Tommaso M, Severi FM, et al
. Maternal anemia effects during pregnancy on male and female fetuses: Are there any differences? J Matern Fetal Neonatal Med 2017;30:1704-8.
Pal A. Role of copper and selenium in reproductive biology: A Brief Update. Biochem Pharmacol 2015;4:5.
Zhang X, Lv F, Tang J. Shear wave elastography (SWE) is reliable method for testicular spermatogenesis evaluation after torsion. Int J Clin Exp Med 2015;8:7089-97.
Eroglu M, Sahin S, Durukan B, Ozakpinar OB, Erdinc N, Turkgeldi L, et al.
Blood serum and seminal plasma selenium, total antioxidant capacity and coenzyme q10 levels in relation to semen parameters in men with idiopathic infertility. Biol Trace Elem Res 2014;159:46-51.
Lukusa K, Lehloenya KC. Selenium supplementation improves testicular characteristics and semen quality of Saanen bucks. Small Rum Res 2017;151:52-8.
O'Hara L, Smith LB. Androgen receptor roles in spermatogenesis and infertility. Best Pract Res Clin Endocrinol Metab 2015;29:595-605.
Di Bona KR, Xu Y, Gray M, Fair D, Hayles H, Milad L, et al.
Short- and long-term effects of prenatal exposure to iron oxide nanoparticles: Influence of surface charge and dose on developmental and reproductive toxicity. Int J Mol Sci 2015;16:30251-68.
Azman M, Senol N. The prevention of the damages of iron (Fe) and zinc (Zn) with the juglone (5-Hydroxy-1, 4-Naphthoquinone) antioxidant activity in the testicular tissue of rats. Fresenius Environ Bull 2016;25:5830-41.
Bano I, Malhi M, Soomro SA, Kandhro S, Awais M, Baloch S, et al
. Effect of dietary selenium supplementation on morphology and antioxidant status in testes of goat. J Basic Appl Sci 2018;14:53-61.
Chen H, Kang Z, Qiao N, Liu G, Huang K, Wang X, et al.
Chronic copper exposure induces hypospermatogenesis in mice by increasing apoptosis without affecting testosterone secretion. Biol Trace Elem Res 2020;195:472-80.
Gunes S, Hekim GN, Arslan MA, Asci R. Effects of aging on the male reproductive system. J Assist Reprod Genet 2016;33:441-54.
Tvrdá E, Lukáč N, Schneidgenová M, Lukáčová J, Szabó C, Goc Z, et al.
Impact of seminal chemical elements on the oxidative balance in bovine seminal plasma and spermatozoa. J Vet Med 2013;2013:125096.
[Figure 1], [Figure 2], [Figure 3], [Figure 4], [Figure 5], [Figure 6], [Figure 7], [Figure 8], [Figure 9], [Figure 10], [Figure 11], [Figure 12], [Figure 13], [Figure 14], [Figure 15], [Figure 16], [Figure 17], [Figure 18]
[Table 1], [Table 2], [Table 3]