Endocrine pancreas development in growth-retarded human fetuses.

Glucose intolerance in adults born with intrauterine growth retardation (IUGR) may involve peripheral insulin resistance and/or abnormal endocrine pancreas development during fetal life. We quantified insulin-containing cells in deceased human fetuses with IUGR (<10th percentile, n = 21) or normal growth (control fetuses, n = 15). Paraffin-embedded pancreatic tissues from fetuses older than 32 weeks were obtained from two fetopathology departments. Mean gestational age was 36 weeks in both groups. Tissues with lysis and those fetuses with defects, aneuploidy, or genetic abnormalities were excluded. For each subject, six pancreatic sections spaced evenly throughout the organ were immunostained with anti-insulin antibody. Total tissue and insulin-positive areas were measured by computer-assisted quantitative morphometry. Results were expressed in percentages. To evaluate islet morphogenesis, the percentages of beta-cells inside and outside islets were determined. Islet density was similar in the two groups (P = 0.86). The percentage of pancreatic area occupied by beta-cells (beta-cell fraction) was not correlated with gestational age (r = 0.06 and P = 0.97 in IUGR fetuses; r = 0.12 and P = 0.67 in control fetuses) or body weight (r = 0.16 and P = 0.47 in IUGR fetuses; r = 0.24 and P = 0.39 in control fetuses). Mean beta-cell fraction was 2.53% in the IUGR fetuses and 2.86% in the control fetuses (P = 0.47). The percentage of beta-cells located within islets was identical in the two groups (mean 35%). Our data militate against a primary developmental pancreatic abnormality in human IUGR, leaving peripheral insulin resistance as the most likely mechanism of glucose intolerance in adults born with IUGR.     

Role of bone bark during growth in width of tubular bones. A study in human fetuses.

The morphologic features of bone bark, a structure surrounding the distal and proximal ends of long bones, were studied in the distal femur, proximal tibia, and proximal fibula of 77 spontaneously aborted human fetuses varying in gestational age from 10 to 20 weeks. Standard histologic techniques used in addition to in situ immunohistochemical staining allowed the examination of the structure of the bone bark and localization of Types 1, 2, and 3 collagens at different gestational ages. The bone bark was shaped like a cylindrical sheath of bone lamellae of varying thickness. The epiphyseal end of the bone bark, known as the groove of Ranvier, was covered outwardly by a fibrous layer and inwardly by the epiphyseal cartilage and contained mesenchymal cells, chondroblastic precursor cells, and densely packed cells differentiating into osteoblasts. Neither the cell density in the groove nor the thickness of the bone bark were identical circumferentially, indicating an unequal growth in width. In addition, the presence of periosteal apposition and endosteal resorption of the bone bark on one side and of endosteal bone deposition accompanied by periosteal resorption of the bone bark on the opposite side support the concept of a spatial drift of bones. These observations furnish histologic proof that groove and bone bark, although assuring an equal growth in length, contribute to an unequal and eccentric growth in width.     

Growth in width of the metacarpals--an investigation in human fetuses.

The process of growth in width of the human hand during fetal life has never been described. Do metacarpals grow concentrically and separation between the bones occurs through expansion of soft tissues? Or is growth eccentric, a process termed drift by Enlow, a relocation in space of organs? Hands of 10 spontaneously aborted fetuses (age range: between 14.5 and 24 weeks of gestation) were examined paying special attention to the bone bark. A thicker bone bark was taken as an indication of growth in that direction. The thickness of the bone bark was measured at the radial and ulnar sides at the level of the proximal and of the distal physes of the second to fifth metacarpals. A ratio of radial over ulnar bone bark thickness (R/U ratio) was calculated. The third metacarpal grew almost concentrically (R/U ratio 1.12 ± 0.06). The second metacarpal grew in a radial direction (R/U ratio 3.29 ± 0.19) and the fourth and more so the fifth metacarpal grew in an ulnar direction (R/U ratio 0.70 ± 0.04 and 0.42 ± 0.02, respectively). The differences in R/U ratios between every metacarpal were statistically significant for all comparisons P ≤ 0.001. Fetal growth in width of the human metacarpals is eccentric and not concentric. It is concluded that during growth in width the metacarpals move away from the midline of the hand and that growth occurs through eccentric bone apposition rather than through soft tissue expansion. © 2001 Orthopaedic Research Society. Published by Elsevier Science Ltd. All rights reserved.     

Brachial plexus variations in human fetuses

OBJECTIVE: We examined the anatomic variations of the brachial plexus (BP) in human fetuses. METHODS: This study was performed with 200 BPs from spontaneously aborted fetuses without detectable malformations. The plexuses were dissected, and the normal position and/or morphological variations of the BP were determined and photographed. RESULTS: There were no variations in 93 plexuses, and 107 plexuses were observed to have different variations. Morphological variations were observed more frequently among female fetuses and right sides. The BPs were composed mostly of the C5, C6, C7, and C8 nerves and the T1 nerve (71.5%). A prefixed plexus was observed in 25.5% of cases, and a postfixed plexus was observed in 2.5% of cases. In one case (0.5%), the C4 and T2 nerves joined the formation. The inferior trunk was not formed in 9% of cases. The superior trunk was not formed in 1% of cases. In one plexus, the superior trunk was formed by the ventral rami of the C4 and C5 nerves. In one case, the inferior trunk was formed by the ventral rami of the T1 and T2 nerves. Division variations were observed most frequently. There were also variations in the terminal branches, such as the roots of the median nerve joining in the distal part of the arm (8.5%), the axillary nerve being separate from the posterior division of the superior trunk (2.5%), and a connection existing between the median and musculocutaneous nerves (1%). CONCLUSION: Knowledge of BP variations is important for surgeons who perform surgical procedures in the cervical and axillary regions.     

Collagen and elastic fibers in the penis of human fetuses at 28 weeks postconception.

OBJECTIVES: The extracellular matrix is a key element in penile function and pathology, yet little is known of its development. Herein we investigated the morphological organization of collagen and elastin in the corpora cavernosa and tunica albuginea of human fetuses. MATERIAL AND METHODS: The penises from 5 fresh human fetuses at 28 weeks postconception (WPC) were routinely fixed and embedded, and all staining procedures were carried out on paraffin sections. Collagen was evidenced by staining with: (1) Gomori's trichrome; (2) sirius red, followed by observation under polarized light, and (3) an antihuman collagen type-III antibody. Elastin and the whole elastic system were revealed using an antihuman elastin antibody and Weigert's resorcin fucsin, respectively. RESULTS: At this stage of fetal development, the albuginea is formed predominantly by dense bundles of collagen. Near the corpora cavernosa, the presence of type-III collagen was also observed. Weigert staining showed numerous fibers of the elastic system in the albuginea. Type-III collagen was found to be strongly positive in the cavernous trabeculae and in the connective sheath surrounding the central artery. Using Weigert staining and an immunolabeling method with primary antibody against human elastin, we found an important quantity of elastic system fibers in the trabeculae of the corpora cavernosa. CONCLUSION: In fetuses at 28 WPC the albuginea is formed predominantly by dense bundles of collagen. The trabecular structures of the corpora cavernosa present a significant quantity of type-III collagen and elastic system fibers.     

Allometry and morphometry of the inguinal region in human fetuses

Background  

The aim of this study was to assess the growth rate of the inguinal region in Brazilian fetuses, irrespective of the knowledge of ethnic genetic background.     

Postnatal human lung growth.

Standard morphometric methods were applied to the lungs of 36 boys and 20 girls aged from 6 weeks to 14 years, dying as a result of trauma or after short illnesses. Individual lung units, alveolar dimensions, and number of alveoli per unit area and volume did not differ between boys and girls, but boys had bigger lungs than girls for the same stature. This resulted in a larger total number of alveoli and a larger aveolar surface area in boys than in girls for a given age and stature. There may be more respiratory bronchioles in boys than girls. There was rapid alveolar multiplication during the first two years of life and alveolar dimensions and number of alveoli per unit area and volume did not change much during this period. There was little or no increase in the total number of alveoli after the age of 2 years but the data are hard to interpret. There is a wide scatter of the total number of alveoli in the growing lung, in keeping with the observation that the total number of alveoli is very variable in adults. Prediction data are given for the various morphometric variables studied.     

Prostate gland growth during development is stimulated in both male and female rat fetuses by intrauterine proximity to female fetuses

In rodents, steroid hormones are transported between adjacent fetuses, and male or female fetuses that develop in utero between female fetuses (2F males or 2F females) have higher serum levels of estradiol and lower serum levels of testosterone relative to siblings of the same sex that develop between two male fetuses (2M males or 2M females). The present study was prompted by the prior unexpected finding that as adults, 2F male mice have an enlarged prostate, and increased numbers of prostatic androgen receptors relative to 2M males. We examined prostate development in both male and female rat fetuses from different intrauterine positions using computer-assisted, 3-dimensional reconstruction of the urogenital complex. In males, this included the prostate, seminal vesicles and utricle (a remnant of the Müllerian ducts), while in females it included development of prostatic glandular buds. The mean cross-sectional area of developing prostatic epithelial buds, utricle and seminal vesicles was significantly increased in 2F male relative to 2M male fetuses. In female fetuses, prostatic bud development was significantly more likely to occur in 2F (67%) than in 2M (29%) animals. These findings suggest that the transport of a small supplement of estrogen from adjacent female fetuses enhances androgen-dependent accessory organ development. We also found that mRNAs encoding receptors for both estrogen and androgen were located in the mesenchyme of the developing male prostate. The localization of estrogen and androgen receptor mRNA in this region further suggests that the mesenchymal induction of prostatic epithelial growth involves both hormones. The cranial dorsolateral prostatic buds exhibited the greatest enlargement in 2F males. This region of the developing prostate in rats is comparable (that is the embryonic homologue) to the region exhibiting benign prostatic hyperplasia (BPH) during aging in men. We propose that the potential for pathological regrowth of the prostate during aging is imprinted by estradiol during fetal development.     

Histochemical and immunohistochemical study of the glomerular development in human fetuses

Few studies exist that establish the normal morphological patterns of glomerular development, though this is one of the organs that continues to evolve morphologically during the postnatal period up to 4 weeks after birth. In our study one kidney from each autopsy of a total of 86 autopsies was analyzed [15 weeks to 40 weeks of gestational age (GA)]. We examined the variation in the nephrogenic zone thickness, the area and diameter of the glomerular tuft, the area and diameter of the glomerular capsule, and the immunohistochemical markers, anti-CD31 and anti-CD34 antibodies, which accompany the development of the glomerular microvasculature. Periodic acid–methenamine silver (PAMS) stain was used for the morphological and morphometrical analyses, and it was particularly useful in fetuses in which autolysis had occurred. The length of the nephrogenic zone (NZ) decreased with the increase of the GA (P < 0.001) according to the formula: . The areas of the Bowman capsule (P < 0.0001), the capillary tuft (P < 0.0001), and the capillary tuft diameter (P = 0.00393) of the intermediary glomeruli increased with the advance of GA, with a positive significant correlation. The same parameters of the juxtamedullary and superficial glomeruli had no correlation with the advance of GA. The cells of the primary structures in the “S” shape of the primitive nephrons were negative for CD31 and CD34. Staining for both antibodies was found, for all GAs, in the endothelial cells of the mature glomeruli tuft and in the renal interstitial vessels. The data obtained in this work contribute to the evaluation of renal maturity in autopsied fetuses and are particularly important in fetuses when autolysis has occurred, to which the parameters used in this study can also be applied. The establishment of normal morphometric and immunohistochemical parameters for the evaluation of renal maturity increases the diagnostic precision of renal pathological alterations in aborted material and perinatal autopsy.     

Congenital absence of the testis in human fetuses and in cryptorchid patients

BACKGROUND: The aim of the present study is to make a comparative study in human fetuses and in patients with cryptorchidism, analyzing the incidence of a number anomalies of the testes for both populations. METHODS: We studied 326 testes from 163 human fetuses ranging in age from 10 to 35 weeks postconception (WPC) and 133 testes from 101 cryptorchid patients aged from 1 to 15 years old (mean, 6.4 years). The Fisher's exact test was used for comparison. RESULTS: Among 326 fetal testes, 224 (68.7%) were abdominal, 45 (13.8%) were inguinal and 55 (16.8%) were scrotal. In one fetus at 23 WPC, both testes (0.6%) were absent. Of the 133 cryptorchid testes, 17 (12.78%) were abdominal, 92 (69.1%) were inguinal and 24 (18%) were high scrotal. Of the 17 abdominal testes, three (17.6%) were atrophic and two were vanished (11.7%). Of the 92 inguinal testes, one (1.08%) was vanished. Twenty-eight (21%) of the cryptorchid testes were impalpable and among these, 17 were located in the abdomen (60.7%) and 11 (38.2%) in the inguinal region (internal ring). CONCLUSIONS: Testicular agenesis is a very rare anomaly, both in fetuses and patients with cryptorchidism.     

Localization of acid phosphatase activity in testosterone-treated prostatic urethra of human fetuses

Androgens accelerated differentiation of human fetal urethral epithelial cells into secretory prostatic cells in organ culture. These epithelial cells contained supranuclear Golgi complexes in which acid phosphatase enzyme activity could be demonstrated by an electron microscopic histochemical technique. Most of the acid phosphatase activity was localized in lysosomes. Some apical precipitates could also be seen. This apical activity might be in secretory granules. Also, mesenchymal cells contained this enzyme.     

Localization of estrogen receptors in long bones and vertebrae of human fetuses

Summary In order to investigate the possible role of estrogen in the development of cartilage and bone we studied by immunofluorescence immunohistochemistry and autoradiography 26 human embryos and fetuses 7–22 weeks in gestational age associated with pregnancy interrupted for nonmedical reasons. In order to demonstrate the presence of estrogen receptors (ERs) in human fetal cartilage, cryostat sections of long bones and lumbar and thoracic vertebrae were prepared for (1) fluorescent immunocytochemistry using an antiidiotypic monoclonal antibody to antiestradiol receptor monoclonal Ab labeled with fluorescein isothiocyanate (FITC), (2) immunohistochemistry using monoclonal antihuman estradiol receptor antibody, labeled with strept. A-B immunoperoxidose, and (3) autoradiographic localization of estradiol using labeled (3 H) l7 estradiol. In fetuses aged 10 weeks or older, intranuclear and perinuclear localization of ER was demonstrated by all methods, mainly amongst chondrocytes of the proliferating and higher hypertrophic zones of the epiphyses and in the cartilage of vertebral bodies. These data suggest that estrogen acts directly on chondrocytes of human fetuses through an ER-mediated mechanism.     

Bombesin stimulates growth of human gastrinoma.

BACKGROUND. We have previously reported the first establishment and characterization of a functioning human gastrinoma (PT) xenograft. Bombesin, the equivalent of the mammalian gastrin-releasing peptide, has trophic effects on normal and neoplastic tissues of the gastrointestinal tract; the effects of gut hormones on the growth of gastrinoma are not known. The purpose of this study was twofold: (1) to determine the presence of various gut peptides in PT and (2) to determine the effect of bombesin on the growth of PT xenografts. METHODS. PT tumors were examined for expression (mRNA and protein) of various gut peptides by Northern hybridization and immunohistochemistry. In addition, PT xenografts were implanted as 3 mm2 pieces bilaterally subcutaneously in athymic nude mice. Mice were divided into two groups to receive either bombesin (5 micrograms/kg) or saline administered as intraperitoneal injections every 8 hours. Tumor area was measured twice weekly until mice were sacrificed (day 28), when tumor and normal pancreas were removed, weighed, and assayed for DNA and protein content. RESULTS. Both mRNAs and peptides of gastrin and chromogranin A were present in PT tumors. Bombesin significantly stimulated growth of PT tumors from day 18 until mice were sacrificed (day 28). As expected, bombesin stimulated pancreatic growth. CONCLUSIONS. We have demonstrated for the first time that bombesin is a trophic hormone for gastrinoma. The unique cell line PT contains gastrin and chromogranin A and will be a useful model to define the biologic mechanisms controlling the growth of human gastrinomas.     

Changes in growth factor levels in human wound fluid.

BACKGROUND. It has been suggested that platelet-derived growth factor (PDGF) plays a central role in wound healing. Analysis of human wound fluid revealed the presence of PDGF AA (30 kd) and monocyte/macrophage-derived growth factor (MDGF) (12 to 14 kd) in the immediate postoperative period. METHODS. The amount of PDGF AA present was assayed by Western blot analysis. The chemotactic and mitogenic potential of purified wound fluid containing PDGF AA and MDGF was determined on a responsive cell line. The biologic activity of MDGF was assayed with a cell line that is unresponsive to the PDGF AA found in wound fluid. RESULTS. Both the concentration and the biologic activity were highest in the immediate postoperative period and declined to negligible levels by 24 hours after surgery. The chemotactic activity of MDGF was highest in the immediate postoperative period and declined during the first 24 hours in a manner similar to that of the combined PDGF AA and MDGF activity. CONCLUSIONS. These data demonstrate the changing levels of PDGF and MDGF in human wound fluid over time, supporting the cascade model of wound repair. By demonstrating that MDGF acts on cell lines unresponsive to the PDGF AA found in wound fluid, these data suggest that MDGF may also play an important role in wound healing.     

The xenotransplantation of goat and human hematopoietic cells to sheep fetuses

Hematopoietic xenografts were carried out in three experiments using goat fetal liver (44-48 days, experiments I and II) or purified human CD 34+ cells (experiment III) as the donor cells. Recipients were sheep fetuses at 41-47 days of gestation. Goat fetal liver cells were either injected without any pretreatment or stimulated by preincubation in a culturing in goat phytohemagglutinin-stimulated lymphocyte supernatant. Human CD 34+ myeloid progenitor cells were purified from bone marrow by minimacs immunomagnetic purification and cultured in medium supplemented with stem cell factor, IL3, and IL6. Goat-sheep chimerism was assessed by flow cytometry analysis (FCA) of peripheral blood and bone marrow cells using a mouse anti-goat CD 45 monoclonal antibody and by karyotype analysis of peripheral blood from goat/sheep chimeras. Human cell engraftment was assessed by polymerase chain reaction amplification of the human DAX1 gene in blood and bone marrow DNA from sheep which had received human cells. In the three experiments, a mean of 76% (26 of 34) of injected fetuses were born alive without any clinical evidence of graft-versus-host disease. Three lambs were found to be goat/sheep chimeric after flow cytometry analysis (peripheral blood and bone marrow) and karyotype (peripheral blood) analysis. Both tissues continued to express goat cells at 6 or 12 months (last assessment) depending on the experiment. No human chimerism was detected using polymerase chain reaction amplification in peripheral blood and bone marrow of any of the six sheep grafted with human cells. These data and those also obtained on other species (human, pig/sheep) show that it is possible to carry out hematopoietic xenografts using the sheep fetus as recipient provided both donor and recipient fetal cells are processed during the period of tolerance to foreign antigens.     

Neurotensin regulates growth of human pancreatic cancer.

OBJECTIVE: The effect of neurotensin (NT) on in vitro-growth of human pancreatic cancer cells (MIA PaCa-2) was examined. Furthermore, the intracellular signal-transduction pathways by which neurotensin regulates growth of MIA PaCa-2 cells were determined. SUMMARY BACKGROUND DATA: NT is trophic for normal rat pancreas, but the effect of NT on growth of human pancreatic cancer is not known. METHODS: Effects of NT (10(-12) to 10(-6) mol/l) on growth of MIA PaCa-2 cells were determined by both count of cell numbers and 3H-thymidine incorporation. Action of NT on phosphatidylinositol (PI) hydrolysis, cyclic AMP production, and intracellular calcium level were determined by conventional methods. The effects of 8-bromo-cyclic AMP and prostaglandin E2 on cell growth were determined. RESULTS: Low concentrations of NT (10(-12) to 10(-9) mol/l) stimulated growth in a dose-dependent manner, but higher concentrations of NT (10(-8) to 10(-6) mol/l) did not stimulate growth of MIA PaCa-2 cells. NT (10(-12) to 10(-6) mol/l) stimulated PI hydrolysis and increased intracellular calcium levels in a dose-dependent manner. High concentrations of NT (10(-8) to 10(-6) mol/l) stimulated production of cyclic AMP in a dose-dependent manner. 8-bromo-cyclic AMP inhibited growth of MIA PaCa-2 cells; prostaglandin E2 did not affect growth of MIA PaCa-2 cells. CONCLUSIONS: NT stimulates growth of MIA PaCa-2 cells through stimulation of PI hydrolysis and mobilization of calcium. Stimulation of the cyclic AMP pathway by high concentrations of NT abolishes the growth-stimulatory effect of NT that is mediated through PI hydrolysis or calcium mobilization.