The role of androgens in development of the scrotum of the grey short-tailed Brazilian opossum ( Monodelphis domestica)

In eutherian mammals, sex differentiation is initiated by expression of the testis-determining gene on the Y chromosome. Subsequent phenotypic development of the reproductive tract and genitalia depends on the production of hormones by the differentiated testis. In marsupials the mechanisms of phenotypic development may vary from this pattern, as differentiation of the scrotal primordia has been shown to occur before that of the gonad. Thus, the development of the scrotum in the marsupial has been regarded as an androgen-independent process. We have sought to clarify the ontogeny of scrotal development and the appearance of androgen receptor immunoreactivity by examining Monodelphis domesticaembryos/pups from 1 day prior to birth until 2 days after birth. We have also used immunocytochemistry to determine the expression of the key steroidogenic enzyme 3beta-hydroxysteroid dehydrogenase as an indicator of when the developing gonad may be capable of synthesizing androgens. Expression of this enzyme was first detected in the gonads and adrenals of both sexes 1 day prior to birth and before the appearance of scrotal bulges. Androgen receptor immunoreactivity was detected in the scrotal anlagen of male opossum pups as early as 1 day following birth. This finding is significantly earlier than previous reports and coincides with the appearance 1 day after birth of distinct scrotal bulges. Androgen receptor immunoreactivity was also observed in the genital tubercles of male pups, but not female pups, 2 days after birth. These results suggest that androgens may play an important role in the development of the male genitalia at a much earlier stage than that indicated by previously published work and that scrotal development in this species may not be androgen-independent.     

Increased levels of apoptosis in the prefusion neural folds underlie the craniofacial disorder, Treacher Collins syndrome

Treacher Collins syndrome (TCS) is an autosomal dominant disorder of human craniofacial development that results from loss-of-function mutations in the gene TCOF1. Although this gene has been demonstrated to encode the nucleolar phosphoprotein treacle, the developmental mechanism underlying TCS remains elusive, particularly as expression studies have shown that the murine orthologue, Tcof1, is widely expressed. To investigate the molecular pathogenesis of TCS, we replaced exon 1 of Tcof1 with a neomycin-resistance cassette via homologous recombination in embryonic stem cells. Tcof1 heterozygous mice die perinatally as a result of severe craniofacial anomalies that include agenesis of the nasal passages, abnormal development of the maxilla, exencephaly and anophthalmia. These defects arise due to a massive increase in the levels of apoptosis in the prefusion neural folds, which are the site of the highest levels of Tcof1 expression. Our results demonstrate that TCS arises from haploinsufficiency of a protein that plays a crucial role in craniofacial development and indicate that correct dosage of treacle is essential for survival of cephalic neural crest cells.     

The role of androgens in development of the scrotum of the grey short-tailed Brazilian opossum (<Emphasis Type="Italic">Monodelphis domestica</Emphasis>)

In eutherian mammals, sex differentiation is initiated by expression of the testis-determining gene on the Y chromosome. Subsequent phenotypic development of the reproductive tract and genitalia depends on the production of hormones by the differentiated testis. In marsupials the mechanisms of phenotypic development may vary from this pattern, as differentiation of the scrotal primordia has been shown to occur before that of the gonad. Thus, the development of the scrotum in the marsupial has been regarded as an androgen-independent process. We have sought to clarify the ontogeny of scrotal development and the appearance of androgen receptor immunoreactivity by examining Monodelphis domestica embryos/pups from 1 day prior to birth until 2 days after birth. We have also used immunocytochemistry to determine the expression of the key steroidogenic enzyme 3β-hydroxysteroid dehydrogenase as an indicator of when the developing gonad may be capable of synthesizing androgens. Expression of this enzyme was first detected in the gonads and adrenals of both sexes 1 day prior to birth and before the appearance of scrotal bulges. Androgen receptor immunoreactivity was detected in the scrotal anlagen of male opossum pups as early as 1 day following birth. This finding is significantly earlier than previous reports and coincides with the appearance 1 day after birth of distinct scrotal bulges. Androgen receptor immunoreactivity was also observed in the genital tubercles of male pups, but not female pups, 2 days after birth. These results suggest that androgens may play an important role in the development of the male genitalia at a much earlier stage than that indicated by previously published work and that scrotal development in this species may not be androgen-independent.     

Neural crest-specific removal of Zfhx1b in mouse leads to a wide range of neurocristopathies reminiscent of Mowat-Wilson syndrome

Mowat-Wilson syndrome is a recently delineated autosomal dominant developmental anomaly, whereby heterozygous mutations in the ZFHX1B gene cause mental retardation, delayed motor development, epilepsy and a wide spectrum of clinically heterogeneous features, suggestive of neurocristopathies at the cephalic, cardiac and vagal levels. However, our understanding of the etiology of this condition at the cellular level remains vague. This study presents the Zfhx1b protein expression domain in mouse embryos and correlates this with a novel mouse model involving a conditional mutation in the Zfhx1b gene in neural crest precursor cells. These mutant mice display craniofacial and gastrointestinal malformations that show resemblance to those found in human patients with Mowat-Wilson syndrome. In addition to these clinically recognized alterations, we document developmental defects in the heart, melanoblasts and sympathetic and parasympathetic anlagen. The latter observations in our mouse model for Mowat-Wilson suggest a hitherto unknown role for Zfhx1b in the development of these particular neural crest derivatives, which is a set of observations that should be acknowledged in the clinical management of this genetic disorder.     

Developmentally regulated expression of the novel cancer anti-apoptosis gene survivin in human and mouse differentiation.

Inhibitors of programmed cell death (apoptosis) may regulate tissue differentiation and aberrantly promote cell survival in neoplasia. A novel apoptosis inhibitor of the IAP gene family, designated survivin, was recently found in all of the most common human cancers but not in normal, terminally differentiated adult tissues. The expression of survivin in embryonic and fetal development was investigated. Immunohistochemistry and in situ hybridization studies demonstrated strong expression of survivin in several apoptosis-regulated fetal tissues, including the stem cell layer of stratified epithelia, endocrine pancreas, and thymic medulla, with a pattern that did not overlap with that of another apoptosis inhibitor, bcl-2. A sequence-specific antibody to survivin immunoblotted a single approximately 16.5-kd survivin band in human fetal lung, liver, heart, kidney, and gastrointestinal tract. In mouse embryo, prominent and nearly ubiquitous distribution of survivin was found at embryonic day (E)11.5, whereas at E15 to -21, survivin expression was restricted to the distal bronchiolar epithelium of the lung and neural-crest-derived cells, including dorsal root ganglion neurons, hypophysis, and the choroid plexus. These data suggest that expression of survivin in embryonic and fetal development may contribute to tissue homeostasis and differentiation independently of bcl-2. Aberrations of this developmental pathway may result in prominent re-expression of survivin in neoplasia and abnormally prolonged cell viability.     

Prostaglandin transporter expression in mouse brain during development and in response to hypoxia

Prostaglandins (PGs) are bioactive lipid mediators released following brain hypoxic-ischemic injury. Clearance and re-uptake of these prostaglandins occur via a transmembrane prostaglandin transporter (PGT), which exchanges PG for lactate. We used Western blot analyses to examine the PGT developmental profile and its regional distribution as well as changes in transporter expression during chronic hypoxia in the neonatal mouse brain. Microsomal preparations from four brain regions (cortex, hippocampus, cerebellum and brainstem/diencephalon) showed gradual increases in prostaglandin transporter expression in all brain regions examined from postnatal day 1 till day 30. There was a significant regional heterogeneity in the prostaglandin transporter expression with highest expression in the cortex, followed by cerebellum and hippocampus, and least expressed in the brainstem/diencephalon. To further delineate the pattern of prostaglandin transporter expression, separate astrocytic and neuronal microsomal preparations were also examined. In contrast to neurons, which had a robust expression of prostaglandin transporters, astrocytes had very little PGT expression under basal conditions. In response to chronic hypoxia, there was a significant decline in PGT expression in vivo and in neurons in vitro, whereas cultured astrocytes increased their PGT expression. This is the first report on PGT expression in the CNS and our studies suggest that PGTs have 1) a widespread distribution in the CNS; 2) a gradual increase and a differential expression in various regions during brain development; and 3) striking contrast in expression between glia and neurons, especially in response to hypoxia. Since PGTs play a role as prostaglandin-lactate exchangers, we hypothesize that PGTs are important in the CNS during stress such as hypoxia.     

HSP110, caspase-3 and -9 expression in physiological apoptosis and apoptosis induced by in vivo embryonic exposition to all-trans retinoic acid or irradiation during early mouse eye development

Apoptosis is an essential physiological process in embryonic development. In the developing eye of vertebrates, three periods of developmental apoptosis can be distinguished: early, intermediate and later. Within the apoptosis pathway, caspases play a crucial role. It has also been shown that HSP110 may have a potential role in apoptosis. The aim of this research was to study the expression of HSP110, caspase-3 and -9 in physiological, retinoic- or irradiation-induced apoptosis during early eye development. Seven pregnant C57Bl/6J mice received 80 mg kg(-1) of all-trans retinoic acid mixed with sesame oil. Seven pregnant NMRI mice received 2 Gy irradiation at the same gestational day. Control mice of both strains (seven mice of each) were not submitted to any treatment. Embryos were harvested at 3, 6, 12 and 24 h after exposition, fixed, dehydrated and embedded. Coronal sections (5 microm) were made. Slide staining occurred alternatively using anti-caspase-3, anti-caspase-9 and anti-HSP110 immunohistochemistry. HSP110 and caspase-3 expression presented similar topographic and chronological patterns, whereas expression of HSP110 was more precocious in retinoic acid-treated embryos. After retinoic exposure, caspase-3- and HSP110-positive cells were increased in the region of the optic vesicle. By contrast, after irradiation, caspase-3- and HSP110-positive cells were noticeably increased in the optic vesicle, peri-optical mesoderm but less in lens placode. HSP110 was expressed before caspase-3. By contrast, caspase-9 was expressed by a very small number of cells in the optic vesicle either under physiological or under teratogenic conditions. Thus, it seems that activation of caspase-9 is dispensable in early eye developmental apoptosis.     

Cloning and developmental expression analysis of the murine homolog of the spinocerebellar ataxia type 1 gene (Sca1)

Spinocerebellar ataxia type 1 (SCA1) is an autosomal dominant neurodegenerative disorder caused by the expansion of a CAG trinucleotide repeat which encodes glutamine in the novel protein ataxin-1. In order to characterize the developmental expression pattern of SCA1 and to identify putative functional domains in ataxin-1, the murine homolog (Sca1) was isolated. Cloning and characterization of the murine Sca1 gene revealed that the gene organization is similar to that of the human gene. The murine and human ataxin-1 are highly homologous but the CAG repeat is virtually absent in the mouse sequence suggesting that the polyglutamine stretch is not essential for the normal function of ataxin-1 in mice. Cellular and developmental expression of the murine homolog was examined using RNA in situ hybridization. During cerebellar development, there is a transient burst of Sca1 expression at postnatal day 14 when the murine cerebellar cortex becomes physiologically functional. There is also marked expression of Sca1 in mesenchymal cells of the intervertebral discs during development of the spinal column. These results suggest that the normal Sca1 gene, has a role at specific stages of both cerebellar and vertebral column development.      

Cartilage anlagen adapt in response to static deformation

Connective tissue adaptation, including the development of cartilaginous anlagen into bones, is widely believed to be related to dynamic, intermittent load and stress histories. Static stresses, on the other hand, are generally believed deleterious in tissue adaptation. Using serial MRI in a natural human experiment (manipulation and corrective casting of infant clubfoot), we have observed casting produces two effects: (1) the well recognized change in relative positions of the hindfoot anlagen; (2) a newly recognized immediate shape change in the anlagen. These changes seemingly enhance the rate of growth of the anlagen and of the ossific nucleus. The shape change or deformation in the anlagen would occur as a result of alterations in the magnitudes and directions of loading from soft tissue attachments and muscle activity and would necessarily be associated with changes in the stress states within the anlagen and, when present, the ossific nuclei. Given the known role of load and stress history in tissue adaptation, we presume the reduced stress histories influence the enhanced growth rates. These observations contradict some current theories of tissue adaptation since static, rather than dynamic stresses play a crucial role in accelerating the growth and development of anlagen in the infant clubfoot.     

Reassessing the role of p53 in cancer and ageing from an evolutionary perspective

The gene p53 has been fashioned as the guardian of the genome and as prototype of the tumour suppressor gene (TSG) whose function must be inactivated in order for tumours to develop. The ubiquitous expression of truncated p53 protein isoforms, results in "premature ageing" of laboratory mouse strains engineered for expressing such isoforms. These facts have been construed in the argument that p53 evolved in order to protect organisms with renewable tissues from developing cancer yet, because p53 is also an inducer of cellular senescence or apoptosis after extensive DNA damage, it becomes a limiting factor for tissue renewal by depleting tissues from stem/precursor cells thus leading to whole-organism ageing. From that point of view p53 displays antagonist pleiotropy contributing to the establishment of degenerative diseases and ageing. Therefore, tumour suppression becomes a balancing act between cancer prevention and ageing. Nevertheless, here we present current evidence showing that the aforementioned argument is rather inconsistent and unwarranted on evolutionary grounds. The evolutionary perspective indicates that p53 evolved so as to play a subtle but very important role during development while its role as a TSG is only important in animals that are protected from most sources of extrinsic mortality, thus suggesting that p53 was primarily selected for its developmental role and not as a TSG. Therefore no real antagonist pleiotropy can be attached to p53 functions and their relationship with whole-organism ageing might be a laboratory artefact.     

小鼠肺发育中细胞增殖与凋亡及相关调控基因的表达

目的 探讨小鼠肺发育过程细胞增殖与凋亡的变化规律及凋亡相关基因ｐ５３、ｂｃｌ－２ｄ在肺发育中的表达意义。方法 不同发育天数的小鼠肺组织,采用免疫组织化学及ＴＵＮＥＬ法,观察细胞增殖与凋亡及相关基因ｐ５３、ｂｃｌ－２的表达。结果 １．胎１４和１６ｄ肺ＰＣＮＡ阳性细胞数达到高峰;之后ＰＣＮＡ阳性率逐渐下降;生后４到１４ｄ,又恢复低水平的细胞增殖,２．小鼠肺发育过程中出现两个细胞凋亡高峰：即胎１４至１６     

Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and Alzheimer's disease

Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is a ubiquitous enzyme that catalyzes the sixth step of glycolysis and thus, serves to break down glucose for energy production. Beyond the traditional aerobic metabolism of glucose, recent studies have highlighted additional roles played by GAPDH in non-metabolic processes, such as control of gene expression and redox post-translational modifications. Neuroproteomics have revealed high affinity interactions between GAPDH and Alzheimer's disease-associated proteins, including the β-amyloid, β-amyloid precursor protein and tau. This neuronal protein interaction may lead to impairment of the GAPDH glycolytic function in Alzheimer's disease and may be a forerunner of its participation in apoptosis. The present review examines the crucial implication of GAPDH in neurodegenerative processes and clarifies its role in apoptotic cell death.     

Apoptosis and its relation with clinical course in patients with Crimean-Congo hemorrhagic fever

Crimean-Congo hemorrhagic fever (CCHF) is a tick-mediated viral infection. Patients with CCHF may show various clinical presentations. The cause of this difference in the clinical course is not completely understood. Apoptosis is programmed cell death and plays an important role in regulating the immune system. Our knowledge of the role of apoptosis in CCHF disease is limited. We investigated the role of apoptosis and their relationship with the severity of the disease in CCHF. Thus, in 30 patients with CCHF and 30 healthy individuals, we analyzed the serum levels of cytochrome C, apoptotic protease activating factor-1 (Apaf 1), caspase 3, caspase 8, caspase 9, sFas, sFasL, perforin, granzyme B, and CK18 by enzyme-linked immunosorbent assay. This is the first study that research the serum levels of the mentioned apoptosis markers in adult patients with CCHF. We found that the serum levels of sFasL, cytochrome C, Apaf 1, caspase 3, caspase 8, caspase 9, perforin, granzyme B, and M30 were statistically significantly different in the acute phase of the disease compared with healthy individuals and patients in convalescent period. There was no association between the clinical severity of the disease and apoptosis markers. In conclusion, the results of our study suggested that the extrinsic and intrinsic apoptosis pathway play an important role in CCHF.     

Apoptosis signaling pathways in lung diseases

Evidence that apoptosis plays an important role in the pathophysiology of lung diseases has been accumulated. Apoptosis signaling is classically composed of two principle pathways. One is a direct pathway from death receptor ligation to caspase cascade activation and cell death. Death receptor ligation triggers recruitment of the precursor form of caspase-8 to a death-inducing complex, through the adaptor protein FADD, which leads to caspase-8 activation. The other pathway triggered by stimuli such as drugs, radiation, infectious agents and reactive oxygen species is initiated in mitochondria. After cytochrome c is released into the cytosol from the mitochondria, it binds to Apaf1 and ATP, which then activate caspase-9. Recently, endoplasmic reticulum has also been shown to be the organelle to execute apoptosis. Further understanding of molecular mechanisms of apoptosis and its regulation by novel drugs may lead to the development of effective strategies against lung diseases. We overview the signaling pathways of apoptosis and discuss the involvement of apoptosis in the pathophysiology of various lung diseases.