Placentation in the African Elephant (Loxodonta africana).

The female elephant shows a 3-week "follicular phase" to commence her 16-week estrous cycle at the end of which a second surge in pituitary luteinizing hormone (LH) release matures and ovulates an ovarian follicle in association with estrous behavior and mating, whereas the first LH surge at the start of the follicular phase causes luteinization of 3-5 partially developed follicles. The prolonged pregnancy of 22 months is supported by a zonary endotheliochorial placenta which secretes placental lactogen (ePL) from around 40 days of gestation in association with replacement of the lumenal epithelium of the endometrium by trophoblast and the development of large corpora lutea (CLs) in the maternal ovaries from the previously formed luteinized follicles in response to the first LH peak early in the follicular phase. The zonary placenta develops above, rather than within, the endometrium. The elephant placenta secretes neither estrogens nor progestagens throughout gestation, as pregnancy maintenance relies on 5α-dihyroprogesterone and other 5α reduced progestagens secreted by secondary CLs stimulated by ePL and the stromal tissue of the fetal gonads, which become extremely enlarged during the second half of the 22-month pregnancy. In female fetuses, this ovarian enlargement includes the development and subsequent regression of multiple primary and secondary follicles with a consequent substantial decline in primary follicle numbers at birth. During the next 8-9 years of pre-pubertal life, however, oocyte and primary follicle numbers recover to levels near those found in late gestation, which may be evidence of postnatal oogenesis occurring in the elephant.

A Novel Objective Method of Estimating the Age of Mandibles from African Elephants (Loxodonta africana Africana).

The importance of assigning an accurate estimate of age and sex to elephant carcasses found in the wild has increased in recent years with the escalation in levels of poaching throughout Africa. Irregularities identified in current ageing techniques prompted the development of a new method to describe molar progression throughout life. Elephant mandibles (n = 323) were studied and a point near the distal dental alveolus was identified as being most useful in ranking each jaw according to molar progression. These 'Age Reference Lines' were then associated with an age scale based on previous studies and Zimbabwean mandibles of known age. The new ranking produced a single age scale that proved useful for both male and female mandibles up to the maximum lifespan age of 70-75 years. Methods to aid in molar identification and the sexing of found jaws were also identified.

Molecular characterization of adipose tissue in the African elephant (Loxodonta africana).

Adipose tissue (AT) is a dynamic and flexible organ with regulatory roles in physiological functions including metabolism, reproduction and inflammation; secreted adipokines, including leptin, and fatty acids facilitate many of these roles. The African elephant (Loxodonta africana) is experiencing serious challenges to optimal reproduction in captivity. The physiological and molecular basis of this impaired fertility remains unknown. AT production of leptin is a crucial molecular link between nutritional status, adiposity and fertility in many species. We propose that leptin has a similar function in the African elephant. African elephant visceral and subcutaneous adipose tissue (AT) was obtained from both sexes and a range of ages including females with known pregnancy status. RNA was extracted and histological sections created and analyzed by microarray, PCR and immunohistochemistry respectively. Gas-chromatography was used to determine the fatty acid composition of AT. Microarray expression profiling was used to compare gene expression profiles of AT from pre-pubertal versus reproductively competent adult African elephants. This study demonstrates, for the first time, leptin mRNA and protein expression in African elephant AT. The derived protein sequence of the elephant leptin protein was exploited to determine its relationship within the class I helical cytokine superfamily, which indicates that elephant leptin is most closely related to the leptin orthologs of Oryctolagus cuniculus (European rabbit), Lepus oiostolus (woolly hare), and members of the Ochotonidae (Pika). Immunohistological analysis identified considerable leptin staining within the cytoplasm of adipocytes. Significant differences in fatty acid profiles between pregnant and non-pregnant animals were revealed, most notably a reduction in both linoleic and α linoleic acid in pregnant animals. This report forms the basis for future studies to address the effect of nutrient composition and body condition on reproduction in captive and wild elephants.

Growth and development of the ovary and small follicle pool from mid fetal life to pre-puberty in the African elephant (Loxodonta africana).

BACKGROUND: Follicle numbers and developing ovarian morphology, particularly with reference to the presence of interstitial tissue, are intimately linked within the ovary of the African elephant during the period spanning mid-gestation to puberty. These have not been previously quantified in any studies. The collection of 7 sets of elephant fetal ovaries between 11.2 and 20.2 months of gestation, and 29 pairs of prepubertal calf ovaries between 2 months and 9 years of age during routine management off-takes of complete family groups in private conservancies in Zimbabwe provided an opportunity for a detailed study of this period. RESULTS: The changing morphology of the ovary is described as the presumptive cortex and medulla components of the fetal ovary settled into their adult form. Interstitial tissue dominated the ovary in late fetal life and these cells stained strongly for 3ß-hydroxysteroid dehydrogenase. This staining continued postnatally through to 4.5 years of age suggesting continued secretion of progestagens by the ovary during this period. The considerable growth of antral follicles peaked at 28% of ovarian volume at around 16.7 months of fetal age. The numbers of small follicles (primordial, early primary and true primary), counted in the cortex using stereological protocols, revealed fewer small follicles in the ovaries of animals aged 0 to 4.5 years of age than during either late fetal life or prepubertal life. CONCLUSIONS: The small follicle populations of the late-fetal and prepubertal ovaries of the African elephant were described along with the changing morphology of these organs. The changes noted represent a series of events that have been recorded only in the elephant and the giraffe species to date. The expansion of the interstitial tissue of the fetal ovary and its continued presence in early post natal life may well contribute to the control of follicle development in these early years. Further research is required to determine the reasons behind the variation of numbers of small follicles in the ovaries of prepubertal calves.

Skeletal development in the African elephant and ossification timing in placental mammals.

We provide here unique data on elephant skeletal ontogeny. We focus on the sequence of cranial and post-cranial ossification events during growth in the African elephant (Loxodonta africana). Previous analyses on ossification sequences in mammals have focused on monotremes, marsupials, boreoeutherian and xenarthran placentals. Here, we add data on ossification sequences in an afrotherian. We use two different methods to quantify sequence heterochrony: the sequence method and event-paring/Parsimov. Compared with other placentals, elephants show late ossifications of the basicranium, manual and pedal phalanges, and early ossifications of the ischium and metacarpals. Moreover, ossification in elephants starts very early and progresses rapidly. Specifically, the elephant exhibits the same percentage of bones showing an ossification centre at the end of the first third of its gestation period as the mouse and hamster have close to birth. Elephants show a number of features of their ossification patterns that differ from those of other placental mammals. The pattern of the initiation of the ossification evident in the African elephant underscores a possible correlation between the timing of ossification onset and gestation time throughout mammals.

The distribution of small preantral follicles within the ovaries of prepubertal African elephants (Loxodonta africana).

BACKGROUND: Data on the distribution of primordial (single layer of squamous granulosa cells), early primary (some granulosa cells cuboidal) and primary (all granulosa cells cuboidal) follicles, grouped together as small follicles (SF) within the ovary of the elephant is lacking, yet such information is necessary to be able to estimate accurately the total numbers of small follicles in the ovaries of elephant throughout their lifespan. AIM: To determine if the density of SF differs between ovaries, between the surfaces of an ovary, or between the interpolar and intermarginal zones of an ovary. MATERIALS/METHODS: Stereological techniques were employed on 25µm thick histological sections of the ovaries recovered from 12 prepubertal elephant calves aged 2 months to 4.5 years. Cell densities were calculated using the optical brick method and Cavalieri's principle for volume calculation. RESULTS: The density of SF (numbers of SF per unbiased counting frame [UCF]) did not differ between the left (1.11±0.39 (mean±sd)) and right (1.10±0.39) ovaries (P=0.82, n=12), or between the lateral (median 1.24; interquartile range 0.85-1.39) and medial (1.03; 0.76-1.36) surfaces of the ovary (P=0.22, n=24) or among the 5 segments of the ovary between the two poles (P=0.20, n=24). The third of the cortex nearest to the mesovarial margin of the ovary had fewer small follicles per UCF (0.85; 0.51-1.28) than the middle third (1.01; 0.78-1.42; P=0.034), and the third adjacent to the free margin (1.27; 0.79-1.51; P=0.0024), n=24 per group. CONCLUSION: Providing a random sample is taken from the full interpolar and intermarginal dimensions of ovary of a non-pregnant elephant, the density of small follicles throughout the cortex may be accurately measured using stereological techniques applied to one of its surfaces.

Early primary--rather than primordial follicles constitute the main follicular reserve in the African elephant (Loxodonta africana).

Information on the ovarian follicle reserve in the African elephant (Loxodonta africana) is lacking. This study set out to determine the ratios of early preantral follicles and their relative dimensions in the ovaries of 16 African elephant aged 10-34 years. The ovaries were sectioned histologically. Follicles were counted and classified according to expansion of the pre-granulosa cells. Early primary follicles were the most common (75.8%±11.8%), followed by true primary follicles (23.8%±11.8%), whereas primordial follicles were the most rare (<2%). Measurements made on at least 100 early preantral follicles from each animal (n=1464) indicate that growth in oocyte and nuclear diameters started with transition to the true primary stage P<0.01. This, together with the observed ratios between the three types of early preantral follicles suggest that both classical primordial and early primary follicles contribute to the ovarian reserve in the African elephant.