Maternal and Offspring Sugar Consumption Increases Perigonadal Adipose Tissue Hypertrophy and Negatively Affects the Testis Histological Organization in Adult Rats

Sugar intake has been associated with the development of male reproductive pathologies because of the increase and dysfunction in different adipose tissue depots. The establishment of these dysfunctions in the early stages of development is unknown. We evaluated the effect of maternal (pregnancy and lactation) and male offspring (from weaning to adulthood) consumption of 5% sucrose on perigonadal adipose tissue (PAT) and testis in adulthood. Moreover, two rat groups were compared, both including pregnant and lactating females: Control (C—drinking tap water) and sugar (S—consuming 5% sucrose solution). From weaning to adulthood with male offspring, four subgroups were formed: Control Mother → Control and Sugar offspring (CC, CS) and Sugar Mother → Control and Sugar offspring (SC, SS). At 120 postnatal days, the testes and PAT were collected and morphologically described. Furthermore, we quantified the number and cross-sectional area of perigonadal adipocytes and their distribution. We found that the males from SC and SS groups showed high PAT weight (p < 0.005), a high number (p < 0.05), and a relative frequency of large adipocytes (p < 0.05), establishing these results during gestational and lactation stages, and enhancing in adulthood since postnatal diet and its interaction. More macrophages, mast cells, and Leydig cells were observed in the interstitial space of the testis for the CS, SC, and SS groups, concluding that consumption of a high-carbohydrate maternal diet, program hypertrophy processes in adult PAT, developing and enhancing with sugar consumption during postnatal life. Furthermore, they are associated with inflammatory processes within the interstitial space of the testis.


INTRODUCTION
The caloric intake is closely related to diets rich in simple and rapidly assimilated sugars, such as glucose, fructose, and sucrose (Murphy and Johnson, 2003). Although the daily recommendation is 5%-10% of carbohydrates from the total energy intake in adults and children (Organization for Economic Co-operation and Development (OECD), 2017), the worldwide consumption of sugars has increased from 169 to almost 180 million metric tons (Murphy and Johnson, 2003; United States Department of Agriculture (USDA), 2020). This increase in the sugar consumption is present from gestation (Gamba et al., 2019;Casas et al., 2020) and breastfeeding (Zou et al., 2012), continuing throughout childhood (Dubois et al., 2007;De León-Ramírez et al., 2021) and adult life (Kumar et al., 2014). During gestation and breastfeeding, nutrition through the maternal diet plays a critical role (Cervantes-Rodríguez et al., 2014;Nicolás-Toledo et al., 2018;Pedrana et al., 2020), it is already well known that imbalances in the adequate consumption of macronutrients, such as proteins (Pedrana et al., 2020) and carbohydrates (Casas et al., 2020), negatively affect the development and maturation of different organs in later stages (Cervantes-Rodríguez et al., 2014;Nicolás-Toledo et al., 2018;Corona-Quintanilla et al., Online ahead of print) because both the glucose and fructose cross the placenta (Holmberg et al., 1956) and fetal development depends on transport of glucose through the mother's blood (Regnault et al., 2013). These programmed or established changes during the fetal/ embryonic stage that can cause diseases in adulthood are known as the theory of the origin and development of diseases (Barker, 1990).
Both high-calorie diets (Jastrzębska et al., 2014;De León-Ramírez et al., 2021) and a BMI > 25 kg/m 2 (overweight and obesity) (Organization for Economic Co-operation and Development (OECD), 2017; Morales et al., 2014) is associated with male reproductive tract diseases related to some metabolic disorders (metabolic syndrome, insulin resistance, and dyslipidemias) (Nicolás-Toledo et al., 2018;De León-Ramírez et al., 2021). In animal models, high-carbohydrate diets have been established depending on the type and amount of mono, di, or polysaccharide used (Rodríguez-Correa et al., 2020). Also, it has already been described that consumption of these diets during pregnancy and postnatal stages can affect the function of the striated muscles associated with copulation (Corona-Quintanilla et al., Online ahead of print) and the expression of factors related to fat, such as the insulin receptor and the development of spermatogonias (Mao et al., 2018). In addition, postnatal consumption negatively affects testicular histology, injuring the intra and extra tubular epithelium (De León-Ramírez et al., 2021).
In mammals such as humans and rats, metabolic relationship between the testis and white adipose tissue (WAT) that surrounds it, better known as perigonadal adipose tissue (PAT), has not yet been clarified (Bjørndal et al., 2011;Luong et al., 2019) and even less the effect of high-carbohydrate diets on communication between both organs, especially if we consider that one of the main targets of metabolic pathologies associated with reproductive tract is adipose tissue, which is particularly vulnerable to changes in nutrition (Bibee et al., 2011).
Considering that limits of the perimeter of adipocytes observed under a histological slide resemble a polygonal mosaic whose vertices point outwards, like a Poisson-Voronoi diagram, the gamma distribution has been used as a statistical model proposal to characterize the adipocyte size distribution (Ibáñez et al., 2018). In this report, measures associated with the spread and shape of the data distribution under the assumption of gamma distribution are used to characterize the size distribution of perigonadal adipocytes and to determine, based on the theory of the developmental origins of the health and disease, its causal relationship with maternal consumption of sucrose and postweaning in male offspring.
The development of alterations in the male reproductive tract has been associated with sugar consumption and metabolic disorders such as for overweight and obesity (Martini et al., 2010;Sadeghi-Bazargani et al., 2013). The progress of these diseases is characterized by an abnormal or excessive accumulation of lipids in the adipocytes of different deposits of WAT (Cinti et al., 2019), as a result of the consumption of high-calorie diets (França et al., 2020;Oliveira et al., 2020). The PAT is one of its largest fat deposits (Berry et al., 2016) and it is the most important for the testis since its extraction in one or both gonads inhibits spermatogenesis (Chu et al., 2010). Although the joint consumption of carbohydrates and fats during pregnancy and postnatal life has adverse effects on the development of germ, Sertoli, and Leydig cells in mice (Mao et al., 2018), and the effect on PAT and testicular epithelium specifically by chronic consumption of simple sugars through maternal diet and throughout postnatal life is still unknown. For this, our objective was to determine the effect of 5% sucrose consumption in maternal (gestation and breastfeeding) and postnatal (childhood and adulthood) stages on the testis and PAT in adult male rats. We hypothesize that maternal consumption of sucrose may program the development of PAT hypertrophy, as well as histological alterations in the testes of male offspring; and this programming could be maintained and enhanced with the consumption of the same disaccharide from early childhood (weaning) to adulthood.

Animals
All procedures applied to animals were approved by the Bioethics Committee the Centro Tlaxcala de Biología de la Conducta from Universidad Autónoma de Tlaxcala, following the Mexican guide for animal care (NOM-062-Z00-1999, Mexico). Twelve pregnant female (14-week-old) Wistar rats (Rattus norvegicus), weighing 220-240 g, were housed individually in polypropylene cages (37 cm × 27 cm × 16 cm) and maintained in controlled rooms with temperatures of 23 ± 1°C and a 12-h light-dark cycle (with the light off from 08:00 h to 20:00 h).

Experimental Design
The details of the experimental design used in this study have been published previously (Corona-Quintanilla et al., Online ahead of print). Primiparous female rats were paired with males of proven fertility. The female was individually placed in polypropylene boxes, considering the day of mating as day 0 of gestation. During pregnancy and breastfeeding, the females were randomly assigned into two groups: a control group (C), which drank unadulterated tap water, and an experimental group that was provided a 5% sucrose solution (S) to drink. Postpartum, the litters were adjusted to 8-10 pups and from postnatal day 22-120, and the male offspring were in pairs or trios placed in polypropylene boxes and fed with either tap water or 5% sucrose according to the assigned experimental group: control mother-control offspring (CC), control mother-sucrose offspring (CS), sucrose mother-control offspring (SC), and sucrose mother-sucrose offspring (SS) (Figure 1).

Dietary Protocol
All rats were fed daily with Chow-5001 standard diet (LabDiet 5001-Laboratory Rodent Diet, St. Louis, MO, United States) and tap water ad libitum, and 5% sucrose was diluted in the tap water for the S groups. The conditions of maintenance and care of the animals were previously established (Corona-Quintanilla et al., Online ahead of print). Notably, this amount of sugar added to the diet does not decrease the energy intake of the equivalent to 15% protein, as we have previously reported (Cervantes-Rodríguez et al., 2014;Corona-Quintanilla et al., Online ahead of print), because the consumption of this nutrient can be considered deficient when it is very low (3% and 5%) and moderately low (8% and 12%), or adequate when equal to 15% and 20% (Moro et al., 2021). This study aims to investigate the effect of sugar consumption without the interference of protein deficiency.
Water and food consumption and body weight were measured daily at 9:30 h during pregnancy, breastfeeding, and adulthood (offspring) (Corona-Quintanilla et al., Online ahead of print) to calculate weight gain, consumption of carbohydrates, lipids, and proteins, as well as their respective individual and total caloric intake. These parameters were daily evaluated according to the nutritional description of the Chow 5001 food.

Obtaining Tissues
Before euthanasia, males from 120 postnatal days (PNd) fasted for 12 h. Male rats were deeply over-anesthetized with sodium pentobarbital (50 mg/kg BW) and decapitated using a rodent guillotine device (Harvard Apparatus, Holliston, MA). After decapitation, animals were placed in a supine position and made a longitudinal incision on the abdominal cavity. Both testes with the epididymis and PAT were harvested and weighed (g tissue/100 g BW) (De León-Ramírez et al., 2021).

Morphometric Analysis
At birth, pup weight, body length (from the tip of the nose to base of tail), heat diameter, abdominal diameter, and anogenital distance were measured using calipers. Left testes were removed, fixed, and dehydrated according to the procedure established by De León-Ramírez et al. (2021). They were embedded in Paraplast Plus (Sigma-Aldrich. St. Lois MO, United States), cut at 7 μm, and stained with hematoxylin and FIGURE 1 | Experimental design. Groups of pregnant and breastfeeding females, and from male rats from weaning to adulthood, fed with a standard diet and tap water (W) without (C) and added 5% sucrose (S).

Estimation of Small and Large Adipocyte Proportions
The cutoff points for small, medium, and large adipocytes were defined based on the 25th and 75th percentiles of the relative distribution of the CC group and analysis by non-parametric comparisons.

Statistical Analyses
The sample size in each group was six rats from different litters. Following the normality of the data (with a frequency distribution histogram), the comparison of groups of mothers during pregnancy and breastfeeding was done by Student's t test.
Two-way ANOVA was used to compare data of body weight, weight gain, food intake, water consumption, testicular, and PAT parameters (number, weight, and CSA of seminiferous tubules and adipocytes, as well as adipocytes size-small, medium, and large proportion) in male offspring for all groups by the combined effects of period diet and sucrose consumption. Period diet (maternal consumption: gestation/breastfeeding; postnatal: childhood/puberty/adulthood) was considered as the first independent variable and sugar intake as the second independent variable. Where ANOVA indicated a significant (p < 0.05) effect of treatments, a post hoc test was carried out using the Tuckey correction to determine significant differences.
The CSA distribution of adipocytes and seminiferous tubules was analyzed using frequency histograms and subsequently Fisher's test by X 2 was used to compare the percentage of area according to size bin.
GraphPad Prism Version 6.0 program for Windows was used and all variables measured were expressed as means ± SEM. Significant differences were considered with a p ≤ 0.05 for all cases.

Growth and Nutritional Consumption of Pregnant Mothers
Differences in weight and consumption of water and food are shown in the Table 1. During pregnancy, there were no differences between the C and S groups in the body weight at beginning of gestation nor in weight gain, but 5% sucrose intake affected the weight, showed at the end of pregnancy (Table 1). Throughout breastfeeding, there were no differences in initial, final, or weight gain. During maternal consumption period, females from the S group significantly increased their intake of both sugar water (98%) and total carbohydrates (by~23%). Likewise the consumption of solid food (lipids and proteins) decreased2 3%; (Table 1). Respect to the protein consumption, it is important to mention that the caloric intake in the S group of dams was equivalent to 18.5%, while 24.1% was for the control group.

Morphometric Measurements of Pups at Birth
Male offspring from dams who consumed sucrose during pregnancy and breastfeeding did not show effects on their initial or final breastfeeding weight, length, head, and abdominal circumference, or anogenital distance ( Table 2).

Growth and Nutritional Consumption of Males During Postnatal Stage
The sucrose intake did not modify the body weight (g) at 22 PNd (CC: 57 ± 2.2, CS: 64 ± 2.8, SC: 58 ± 3.2, and SS: 60 ± 4.1) at any life period (maternal consumption diet: p = 0.5732; postnatal diet: p = 0.1960; and interaction: p = 0.4807). There was an increase in the weight gain after 10 weeks in those groups that consumed sucrose at some period of their development, compared to the CC group ( Figure 2A). This increase in the weight varied accordingly to the period of life in which sucrose was intake. Significant differences were found from 12 to 17 weeks since prenatal intake (CC vs. SC, Figure 2B), during the 7th week and from 12 to 17 weeks since postnatal consumption (CC vs. CS, Figure 2C), and from 13 to 17 weeks since consumption interaction in both periods (CS vs. SC and CC vs. SS; Figures 2D,E).
The postnatal sucrose intake had effect on both the body weight at 120 PNd (p ≤ 0.0001) and weight gain (p ≤ 0.0001) (Figures 2A,C). However, there was a significant effect by the interaction (Figures 2D,E) of maternal and postnatal diets (final weight: p = 0.0005 and weight gain: p = 0.0160). The effect of the postnatal diet was enhanced by the maternal diet (final weight, p = 0.0596 and weight gain, p = 0.0740).
All data related to water and food intake in male groups are presented in the Table 2.
The initial, final, and total food consumption at the postnatal period was significantly decreased only in the groups with 5% sucrose postweaning intake. At the beginning of this study, there were no differences in the food intake between the groups. However, at the final of treatment, the food intake (CC vs. CS p ≤ 0.005; CC vs. SS p ≤ 0.005; CS vs. SC p ≤ 0.05; and SC vs. SS p ≤ 0.05) and total consumption (CC vs. CS p ≤ 0.005; CC vs. SC p ≤ 0.005; and SC vs. SS p ≤ 0.05) were different.
The initial water consumption was not affected by the maternal or postnatal sucrose intake, nor by the interaction ( Table 2). However, in the postnatal stage, the final (CC vs. CS p ≤ 0.005; CC vs. SS p ≤ 0.005; CS vs. SC p ≤ 0.005; and SC vs. SS p ≤ 0.005) and total (CC vs. CS p ≤ 0.005; CC vs. SS p ≤ 0.005; CS vs. SC p ≤ 0.005; and SC vs. SS p ≤ 0.005) water consumption was significantly increased by 5% sucrose intake. 2 | Weight, morphometric measurements, and water and food intake obtained at the birth, breastfeeding (from 1 to 21 PNd), and postnatal stage (22 to 120 PNd) of male offspring that consumed sucrose at 5% (CS and SS) and simple (CC and SC) water and whose mothers during gestation and breastfeeding consumed the same diets. The effect of diets during the pre (MD) and postnatal (PDN) periods and their interaction are shown. Data related to nutritional intake are presented in the Table 3. The decrease in the consumption of both proteins and lipids and their respective energy calculation in the CS and SS groups were due to consumed postnatal sucrose with extremely significant effect (corresponding to 49% of the variance in said variables), as well as significant differences were observed between groups. In the CS, SC, and SS groups, the decrease in the lipids and proteins was 19%, 4.5%, and 20%, respectively. Specifically, the equivalent of protein caloric intake in these groups ranged between 19% and 23% (CS: 19.5%; SC: 23.0%; and SS: 19.2%), compared to the CC group which was 24.1%.

Pups
The increase in the carbohydrate consumption was 15% and 12% in the CS and SS groups compared to the CC group. This increase (38.7% of the total variance) was associated with postnatal carbohydrate consumption and its energy calculation. A significant difference between groups was found only between the CS and SC groups. Both the total caloric intake and hematocrit level in adult males for all groups were unaffected by maternal and postnatal diets, or their interaction.

Morphometric Analysis of the Testis
Testis variables such as the relative weight, CSA, and number of seminiferous tubules were unaffected by maternal and postnatal diets or their interaction (Table 3). Relative distribution of the CSA of the seminiferous tubules was carried out (Figure 3). The seminiferous tubules had CSA ranging from 20 to 120 μm 2 , FIGURE 2 | Weekly weight gain (A) of CC (white circle and solid line), CS (black circle with dotted line), SC (black square with dotted line), and SS (white square with dotted line) male groups from weaning at 120 PNd. Comparisons per week between groups of males that presented differences due to maternal (B) or postnatal diet (C) and interaction (D,E). Mean values ±SEM (n = 6) are shown. Data were analyzed with a two-way ANOVA and Tukey post hoc. *p < 0.05, **p < 0.01, ***p < 0.001, and ****p < 0.0001.
Frontiers in Cell and Developmental Biology | www.frontiersin.org June 2022 | Volume 10 | Article 893099 3 | Sum of total consumption and energy intake of proteins, lipids, and carbohydrates from 4 to 17 weeks, and parameters testis in the groups of males that consumed sucrose at 5% (CS and SS) and simple (CC and SC) water during the postnatal stage (22 to 120 PNd) and whose mothers during pregnancy were fed the same diets. The effect of diets during the maternal (MD) and postnatal (PND) periods and their interaction are shown.  finding the highest percentage (more than 50%) among 40-59 μm 2 , and presenting a Gaussian distribution around this interval. However, there were no differences between them (Figure 3).
No differences in the cell arrangement, typical presence of Sertoli cells in the basal area, germ cells (spermatogonia, spermatocytes, spermatids, and sperm), tunica propria (peritubular tissue of myoid cells) of each tubule, Leydig cells, and other populations of interstitial cells were found between groups ( Figures 4A-H). However, a high number of interstitial cells, macrophages, and mast cells were observed in those groups that consumed sucrose at some stage of their development ( Figures 4A-H).

Morphometric Analysis of Perigonadal Adipose Tissue
The relative weight of the PAT ( Figure 5A) for the CS (0.6 g ± 0.1) and SS (0.7 g ± 0.1) groups was significantly increased in comparison with the CC (0.4 g ± 0.1) and SC (0.4 g ± 0.1) groups, having 20.2% of variation to the effect of 5% sucrose postnatal consumption (p = 0.0044). Neither the maternal diet (p = 0.604) nor the interaction (p = 0.8774) of diets between both periods had any significant effect on this variable. The number of adipocytes ( Figure 5B) was significantly increased by the maternal diet (p = 0.0189), interaction (p = 0.0214), and postnatal diet (p = 0.0528).

Frequency Distribution of Perigonadal Adipocytes
The relative frequency distribution of adipocytes with intervals of 200 units in the four groups of males ( Figure 6A) presented a Gaussian-type distribution, loaded toward the extreme left, with a high proportion of adipocytes with CSA smaller than 1051 μm 2 .
Only groups that consumed sucrose at some period of their life presented adipocyte proportions with CSA greater than 3,600 μm 2 ( Figures 6B-D), especially the SC and SS groups ( Figures 6C,D), whose proportion of adipocytes with CSA of 4,800-9,000 μm 2 was constant in almost all established intervals.
Comparisons with Fisher's test by X 2 between groups with intervals of 200 units were done, resulting in significant differences in adipocytes with CSA from 1001 to 1200 and 1201 to 1400 by postnatal diet (CC vs. CS, p = 0.0400). However, to properly distinguish the effect of diet in the two periods, one more comparison (data not plotted) was done with frequency intervals of 400 units. Resulting in a significant decrease in the proportion of CSA adipocytes from 801 to 1200 μm 2 (p = 0.0281) by maternal diet (CC vs. SC) and in those from 1,201 to 1,600 μm 2 (p = 0.0414) by postnatal diet (CC vs. CS). To corroborate these results, two comparisons were done with frequency intervals of 600 and 1200 μm 2 (histograms are not FIGURE 5 | Relative weight of TAP (A), number (B), and cross-sectional area (CSA) (C) of adipocytes of males that consumed 5% sucrose (gray box) and tap water (black box) during the postnatal period and whose mothers during pregnancy and breastfeeding were fed with same diets. Data were analyzed with a two-way ANOVA and Tukey post hoc. Significant differences are shown. *p < 0.05, **p < 0.01, and ***p < 0.001.

Estimation of Small and Large Perigonadal Adipocyte Proportions
The cutoff points for small and large adipocytes were defined based on the 25th and 75th percentiles of the CSA from CC group distribution ( Figure 6A), which were 1,051 μm 2 for small adipocytes and 1,789 μm 2 for large ones.
When the relative percentage of small, medium, and large adipocytes between groups was compared (Figure 7), only significant differences in the proportions of small and large of the CC group with those groups that consumed sucrose at some periods of their life were found. In both comparisons, sucrose consumption affected in some way the development of large adipocytes, resulting in a significant decrease in the relative percentage of frequency of small adipocytes (CC: 32.6%, CS: 16.1%, SC: 15.4%, and SS: 17.3%) and an increase in large adipocytes (CC: 29.5%, CS: 54.0%, SC: 52.6%, and SS: 51.0%). The highest percentages in the adipocyte CSA were located between intervals (bin 200) of 1,200-1,800 μm 2 (overall) in all groups (CC: 37.7%, CS: 29.8%, SC: 31.9%, and SS: 32.5%) without differences between them (Figure 7).
These differences between the proportion of small and large adipocytes were observed when drawing with different colors the classification of the CSA adipocyte in the photomicrographs taken at ×10, presenting a high proportion of small adipocytes in the CC group ( Figure 6A) compared to the groups that consumed sucrose at some stage of their life, since the CS ( Figure 6B), SC ( Figure 6C), and SS ( Figure 6D) groups presented a greater number of large than small adipocytes.
The adipocytes of the CC group were observed to be proportionally amorphous, asymmetric, and smaller ( Figure 6A) compared to those of the CS, SC, and SS groups ( Figures 6B-D), which were observed to be more rounded, symmetric, and larger.

DISCUSSION
Excessive sugar consumption during gestation is positively associated with maternal weight gain at the end of this period (Renault et al., 2015). Findings of the present study confirm that mothers who consumed sucrose throughout gestation increased the consumption of water with sucrose and in consequence the total carbohydrates amount, resulting in a significant increase in The color of bars in graphics corresponds with the outline drawn of adipocytes. Blue (small, ≤1,051 μm 2 ), red (medium, from 1,052 to 1,788 μm 2 ), yellow (large, ≥1,789 μm 2 ), and purple (those with a CSA greater than 4,000 μm 2 ).
Frontiers in Cell and Developmental Biology | www.frontiersin.org June 2022 | Volume 10 | Article 893099 the weight at the end of this period. This is because carbohydrate consumption consequently leads to an accumulation of triglycerides in different WAT deposits, which increase their size (hypertrophy) (Morigny et al., 2021). At birth, the consumption of sucrose increased slightly (not significant data) the body length of the offspring. This is likely associated with the glycemic index, which is related to neonatal central adiposity measured by the ratio waist-length in humans (Horan et al., 2014). For its part, birth weight was unaffected by sucrose consumption, as it has been reported in other studies with a moderate maternal nutritional excess (Petherick et al., 2014). Both in pregnant mothers and adult male groups, there were no differences along the growth trajectory. But a positive relationship was found between the increase in the body weight and weight gain at the end of treatments because the postnatal diet and its interaction with maternal diet, as it had been reported in mice (Mao et al., 2018). Highlighting that in our model, the weight increase begins after the 3rd week of sucrose consumption. This effect on the body weight has been previously described with high carbohydrates diets (Adekunbi et al., 2016;Mao et al., 2018;De León-Ramírez et al., 2021), indicating that the consumption of these sugars (regardless of whether it is mono or disaccharide) is associated with the regulation of solid food intake to adapt energy provided by the increase in the consumption of sugar water (Sheludiakova et al., 2012;Cervantes-Rodríguez et al., 2014).
Respect to the protein consumption, despite the fact that their caloric intake was reduced, it did not imply a deficiency in the consumption of this nutrient, since both in pregnant females that consumed sucrose (S, 18.5%), as in the adult offspring that postnatally consumed it (CS, 19.4%; SS, 19.2%) and in those males control that came from mothers that consumed during pregnancy and breastfeeding (SC, 23%). This intake ranged from 15% to 20% of total caloric intake, which is considered within the optimal range, even between 20% and 25%, which is desirable for these animals (Moro et al., 2021).
We did not observe changes in the number of seminiferous tubules counted, since, as already described, the rat testicle is composed of approximately 20 seminiferous tubules (Clermont and Huckins, 1961), with a spatial distribution that depends on its connection with the rete testis (Nakata et al., 2015). Also, because the complete development of the number of these tubules occurs during the gestational stage, changes in this variable have been reported when interventions by mutagenic, teratogenic, or carcinogenic agents have been carried out at this stage (Tirpák et al., 2021).
Blood-testicular barrier (HTB) maintains the balance in the testicular environment, promoting a healthy and complete development of cell types in these organs (Banks et al., 1999). However, it has been reported that high concentrations of sugar ingested in postnatal stages (Mao et al., 2018;De León-Ramírez et al., 2021) can modify apoptotic processes in the germ and Sertoli cells (Mao et al., 2018), as well as the CSA, epithelial area (germinative, luminal, interstitial), and testicular weight (De León-Ramírez et al., 2021). Probably due to significant increases in the leptin levels that could cross HTB and modify the communication with testicular triglycerides or with important hormones such as testosterone (Mao et al., 2018;De León-Ramírez et al., 2021). In the same context, Despite not finding structural changes in the morphology of the main cell types intra (Sertoli cells, spermatogonia, spermatocytes, spermatids, and spermatozoa) and extra tubular (Leydig cells), we observed a high proportion of macrophages and interstitial mast cells in groups that consumed sucrose maternal and postnatally, which can be related to a pro-inflammatory status with the possibility of affecting testosterone synthesis and its consequent metabolic and cellular interactions (Harris et al., 2016;Ye et al., 2021), including its communication with WAT deposits like the perigonadal.
WAT had hypertrophy and hyperplasia, and the balance between these two mechanisms is an important factor determining the ultimate outcome in lipid storage homeostasis. For this reason, the determination of the size and number of adipocytes in different WAT deposits has focused on finding new and better strategies for their evaluation and understanding of the involved metabolic dynamics (Ibáñez et al., 2018).
Although in our study a Gaussian-type distribution was determined for adipocytes size, it also present characteristics that could be considered a gamma distribution, which has been used to evaluate the size adipocytes distribution from offspring due to the effect of obesogenic maternal diet (Ibáñez et al., 2018).
As it has been reported for the WAT, an increase in the content of triglycerides may be associated with hypertrophy and/ or hyperplasia of adipocytes of the PAT (Morigny et al., 2021). We found that both processes were triggered and increased by the high postweaning sucrose intake and programmed by the same diet during pregnancy and lactation. A decrease in the relative percentage of small adipocytes was observed in adulthood, leading to a significant increase in the number and the FIGURE 7 | Relative frequency of CSA adipocytes according to their size category for the CC (white bars), CS (diagonal lines bars), SC (vertical lines bars), and SS (black bars) groups. Means ± SEM. Data were analyzed with a two-way ANOVA and Tukey post hoc. Significant differences among intervals are shown. *p < 0.05, **p < 0.01, and ***p < 0.001.

DATA AVAILABILITY STATEMENT
The raw data supporting the conclusion of this article will be made available by the authors, without undue reservation.

ETHICS STATEMENT
The animal study was reviewed and approved by the Comité de Ética del Centro Tlaxcala de Biología de la Conducta del Universidad Autónoma de Tlaxcala.