Immunoelectron microscopy provides evidence for the presence of mitochondrial heat shock 10-kDa protein (chaperonin 10) in red blood cells and a variety of secretory granules

Hsp10 (10-kDa heat shock protein, also known as chaperonin 10 or Cpn10) is a co-chaperone for Hsp60 in the protein folding process. This protein has also been shown to be identical to the early pregnancy factor, which is an immunosuppressive growth factor found in maternal serum. In this study we have used immunogold electron microscopy to study the subcellular localization of Hsp10 in rat tissues sections embedded in LR Gold resin employing polyclonal antibodies raised against different regions of human Hsp10. In all rat tissues examined including liver, heart, pancreas, kidney, anterior pituitary, salivary gland, thyroid, and adrenal gland, antibodies to Hsp10 showed strong labeling of mitochondria. However, in a number of tissues, in addition to the mitochondrial labeling, strong and highly specific labeling with the Hsp10 antibodies was also observed in several extramitochondrial compartments. These sites included zymogen granules in pancreatic acinar cells, growth hormone granules in anterior pituitary, and secretory granules in PP pancreatic islet cells. Additionally, the mature red blood cells which lack mitochondria, also showed strong reactivity with the Hsp10 antibodies. The observed labeling with the Hsp10 antibodies, both within mitochondria as well as in other compartments/cells, was abolished upon omission of the primary antibodies or upon preadsorption of the primary antibodies with the purified recombinant human Hsp10. These results provide evidence that similar to a number of other recently described mitochondrial proteins (viz., Hsp60, tumor necrosis factor receptor-associated protein- 1, P32 (gC1q-R) protein, and cytochrome c), Hsp10 is also found at a variety of specific extramitochondrial sites in normal rat tissue. These results raise important questions as to how these mitochondrial proteins are translocated to other compartments and their possible function(s) at these sites. The presence of these proteins at extramitochondrial sites in normal tissues has important implications concerning the role of mitochondria in apoptosis and genetic diseases.

Induction of birth in the bandicoot (Isoodon macrourus) with prostaglandin and oxytocin

As in eutherians, maturation of the fetal pituitary and adrenal glands together with an increase in prostaglandin and mesotocin or oxytocin production initiates birth in marsupials. in this study, prostaglandin (Lutalyse) or oxytocin (Syntocinon) were administered to pregnant bandicoots at 05:00 h on the calculated day of birth and the resultant effects were filmed for analysis. The administration of prostaglandin caused the bandicoot to adopt the birth position several minutes after injection (n = 2). However, the bandicoot did not give birth for several hours. Birth occurred at a similar time of day to that observed for untreated bandicoots (n = 7), between 08:00 h and 12:00 h. After an injection of oxytocin, the bandicoot assumed the birth position and birth occurred within several minutes. The young were alive while still connected to their allantoic stalks. However, they were unable to attach to the teats and did not survive (n = 4). The induced young were the colour of venous blood and died soon after the umbilicus was separated, indicating that the cardiopulmonary system of these neonates was underdeveloped and inadequate to maintain life. The results from this study demonstrate that prostaglandin is required to prepare the bandicoot for birth, and mesotocin is required for contraction of the uterus and for birth to occur.

Hand-assisted laparoscopic adrenalectomy: An alternative minimal invasive surgical technique for the adrenal gland

Background: Presently the surgical approach to the adrenal gland is in a state of flux. While the traditional approach to the adrenal gland has been the open transabdominal technique, more recently laparoscopic approaches, particularly via the transabdominal route, have increasingly been utilized. However, laparoscopic intervention for the adrenal gland can be problematic in certain circumstances, particularly for large adrenal masses and in instances of adrenal malignancies. Methods: In this report we describe the use of hand-assisted laparoscopic adrenalectomy as an alternative minimal invasive surgical approach to the adrenal gland. Hand-assisted laparoscopic adrenalectomy using the HandPort system (Smith & Nephew, Sydney, Australia) was undertaken in three patients requiring adrenalectomy for mass lesions including one patient with Conn's syndrome. Results: In all three cases, surgery proceeded promptly and uneventfully. In the present paper, the details of the technique of hand-assisted adrenalectomy are described. This is the first report in the world literature of this new technique for the adrenal gland. Conclusions: Hand-assisted laparoscopic adrenalectomy is an easily performed technique, which can be completed within a short operative time span and which has the advantage of providing intraoperative tactile localization for the adrenal gland. It may be particularly applicable for large adrenal tumours, yet only involves the performance of a small abdominal incision. Postoperative recovery is comparable with that reported for the laparoscopic-only technique. Hand-assisted adrenalectomy is a new technique which has great potential and which warrants further evaluation.

Prolactin-releasing peptide in ewe: cDNA cloning, mRNA distribution and effects on prolactin secretion in vitro and in vivo

RT-PCR followed by 5'- and 3'- rapid amplification of cDNA ends was used to clone and sequence ovine prolactin-releasing peptide (PrRP). The cDNA was characterised by short 5'- and 3'-untranslated regions and a GC-rich (71%) coding region. The nucleotide and deduced amino acid sequences for the coding region showed 95.6 and 94.9% identity with bovine PrRP but the amino acid sequence of PrRP31 was conserved between these species. Northern blot analysis and RT-PCR showed that, as in the rat, the peptide was more abundantly expressed in the brainstem than the hypothalamus. However, in the ovine hypothalamus, PrRP mRNA expression was more widespread than in the rat, with expression detected in both rostral and caudal parts of the mediobasal hypothalamus. The effects of synthetic ovine PrRP on prolactin secretion both in vitro and in vivo were also examined. In primary cultures of sheep pituitary cells, PrRP significantly (P

Distribution of two splice variants of the glutamate transporter GLT-1 in rat brain and pituitary

We have performed immunocytochemistry on rat brains using a highly specific antiserum directed against the originally described form of the glutamate transporter GLT-1 (referred to hereafter as GLT-1alpha), and another against a C-terminal splice variant of this protein, GLT-1B. Both forms of GLT-1 were abundant in rat brain, especially in regions such as the hippocampus and cerebral cortex, and macroscopic examination of sections suggested that both forms were generally regionally coexistent. However, disparities were evident; GLT-1alpha was present in the intermediate lobe of the pituitary gland, whereas GLT-1B was absent. Similar marked disparities were also noted in the external capsule, where GLT1A labeling was abundant but GLT-1B was only occasionally encountered. Conversely, GLT-1B was more extensively distributed, relative to GLT-1alpha, in areas such as the deep cerebellar nuclei. In most regions, such as the olfactory bulbs, both splice variants were present but differences were evident in their distribution. In cerebral cortex, patches were evident where GLT-1B was absent, whereas no such patches were evident for GLT-1alpha. At high resolution, other discrepancies were evident; double-labeling of areas such as hippocampus indicated that the. two splice variants may either be differentially expressed by closely apposed glial elements or that the two splice variants may be differentially targeted to distinct membrane domains of individual glial cells. (C) 2002 Wiley-Liss, Inc.

Localization of taurine transporters, taurine, and H-3 taurine accumulation in the rat retina, pituitary, and brain

The nervous system contains an abundance of taurine, a neuroactive sulfonic acid. Antibodies were generated against two cloned high-affinity taurine transporters, referred to in this study as TAUT-1 and TAUT-2. The distribution of such was compared with the distribution of taurine in the rat brain, pituitary, and retina. The cellular pattern of [H-3] taurine uptake in brain slices, pituitary slices, and retinas was examined by autoradiography. TAUT-2 was predominantly associated with glial cells, including the Bergmann glial cells of the cerebellum and astrocytes in brain areas such as hippocampus. Low-level labeling for TAUT-2 was also observed in some neurones such as CA1 pyramidal cells. TAUT-1 distribution was more limited; in the posterior pituitary TAUT-1 was associated with the pituicytes but was absent from glial cells in the intermediate and anterior lobes. Conversely, in the brain TAUT-1 was associated with cerebellar Purkinje cells and, in the retina, with photoreceptors and bipolar cells. Our data suggest that intracellular taurine levels in glial cells and neurons may be regulated in part by specific high-affinity taurine transporters. The heterogeneous distribution of taurine and its transporters in the brain does not reconcile well with the possibility that taurine acts solely as a ubiquitous osmolyte in nervous tissues. (C) 2002 Wiley-Liss, Inc.

Role of PGF(2 alpha) and oxytocin in parturition in the brushtail possum (Trichosurus vulpecula)

Maturation of the fetal pituitary and adrenal glands allows the secretion of cortisol, which in turn leads to an increase in prostaglandin and mesotocin production. The production of prostaglandin and mesotocin results in an increase in uterine contractions and initiates birth in marsupials. The major metabolite of PGF(2alpha), 13,14-dihydro-15-keto-prostaglandin F-2alpha (PGFM), has been found in the plasma of the possum at the time of birth and administration of PGF(2alpha) to female possums induced the adoption of the birth position. Evidence that mesotocin is an integral hormone of birth in the tammar wallaby indicates that both PGF(2alpha) and mesotocin or oxytocin are required for marsupial birth. The presence of PGF(2alpha) receptors in the uterus and corpus luteum of the possum, and the in vitro uterine responsiveness to PGF(2alpha) or oxytocin, were examined. PGF(2alpha) receptors were not observed in possum uteri and the inability of PGF(2alpha) to cause contractions indicates that PGF(2alpha) is not involved directly in contraction of the uterus at parturition. The presence of oxytocin and mesotocin receptors in the uterus of possoms and the ability of oxytocin to induce uterine contraction in vitro supports the view that mesotocin is required for expulsion of the young from the uterus. Low numbers of PGF(2alpha) receptors were found in the possum corpus luteum at birth, indicating an involvement of PGF(2alpha) in regression of the corpus luteum.

Microstructural evolution of the Serido Belt, NE-Brazil: the effect of two tectonic events on development of c-axis preferred orientation in quartz

The polyphase evolution of the Serido Belt (NE-Brazil) includes D, crust formation at 2.3-2.1 Ga, D-2 thrust tectonics at 1.9 Ga and crustal reworking by D-3 strike-slip shear zones at 600 Ma. Microstructural investigations within mylonites associated with D-2 and D-3 events were used to constrain the tectono-thermal evolution of the belt. D-2 shear zones commenced at deeper crustal levels and high amphibolite facies conditions (600-650 degreesC) through grain boundary migration, subgrain rotation and operation of quartz Q-prism slip. Continued shearing and exhumation of the terrain forced the re-equilibration of high-T fabrics and the switching of slip systems from (c)-prism to positive and negative (a)-rhombs. During D-3, enhancement of ductility by dissipation of heat that came from syn-D-3 granites developed wide belts of amphibolite facies mylonites. Continued shearing, uplift and cooling of the region induced D-3 shear zones to act in ductile-brittle regimes, marked by fracturing and development of thinner belts of greenschist facies mylonites. During this event, switching from (a)-prism to a basal slip indicates a thermal path from 600 to 350 degreesC. Therefore, microstructures and quartz c-axis fabrics in polydeformed rocks from the Serido Belt preserve the record of two major events, which includes contrasting deformation mechanisms and thermal paths. (C) 2003 Elsevier Ltd. All rights reserved.

Abnormalities of the PTH-vitamin D axis and bone turnover markers in children, adolescents and adults with cystic fibrosis: comparison with healthy controls

Abnormalities of calcium and vitamin D metabolism in cystic fibrosis (CF) are well documented. We tested the hypothesis that alterations in calcium metabolism are related to vitamin D deficiency, and that bone resorption is increased relative to accretion in patients with CF. Calcitropic hormones, electrolytes, osteocalcin (OC) and bone alkaline phosphatase (BAP), (markers of bone mineralisation), urinary deoxypyridinoline [total (t) Dpd, a marker of bone resorption] and lumbar spine bone mineral density (LS BMD), expressed as a z-score, were measured in 149 (81 M) CF and 141 (61 M) control children aged 5.3-10.99 years, adolescents aged 11-17.99 years and adults aged 18-55.9 years. Data were analysed by multiple regression to adjust for age. In patients, FEV1% predicted and CRP (as disease severity markers), genotype and pancreatic status (PS) were recorded. The distribution of PTH differed between groups (P

Adrenal function in 23 children with paracoccidioidomycosis

Adrenal involvement by Paracoccidioides brasiliensis was described at necropsies and in many clinical studies, but only in adults. Therefore, the aim of this study was to evaluate adrenal function in children with paracoccidioidomycosis. Twenty-three children with the systemic form of paracoccidioidomycosis were evaluated and divided in two Groups: Group A (n = 8) included children before treatment and Group B (n = 15) children after the end of treatment. Plasma cortisol (basal and after ACTH test), ACTH, renin activity, aldosterone, sodium and potassium were measured. They were within normal range in all cases, except for renin activity and aldosterone, which were elevated in some cases. Group A patients showed basal and post-ACTH cortisol levels significantly greater than Group B patients. The results showed that adrenal function was not compromised in these children with paracoccidioidomycosis.

Identification of Paracoccidioides brasiliensis in adrenal glands biopsies of two patients with paracoccidioidomycosis and adrenal insufficiency

The authors report two cases of adrenal insufficiency secondary to infiltration of the adrenal glands by Paracoccidioides brasiliensis. The first patient had been treated for a chronic multifocal form of paracoccidiodomycosis 11 years ago. The diagnosis of the mycosis was done simultaneous with that of the adrenal insufficiency in the second patient. In both patients the diagnosis was done by direct visualization of fungus in adrenal biopsies. They were treated with hormonal supplements and itraconazol by 12 and six months, without relapses during the follow-up period.

DISSEMINATED FUNGAL INFECTION WITH ADRENAL INVOLVEMENT: REPORT OF TWO HIV NEGATIVE BRAZILIAN PATIENTS

Paracoccidioidomycosis and histoplasmosis are systemic fungal infections endemic in Brazil. Disseminated clinical forms are uncommon in immunocompetent individuals. We describe two HIV-negative patients with disseminated fungal infections, paracoccidioidomycosis and histoplasmosis, who were diagnosed by biopsies of suprarenal lesions. Both were treated for a prolonged period with oral antifungal agents, and both showed favorable outcomes.

Aggressive Pituitary Lesion with a Remarkably High Ki-67

Os raros tumores pituitários agressivos são chamados carcinomas quando são detectadas metástases, sejam sistêmicas e/ou em sistema nervoso central. Alguns casos estão associados com superprodução de hormônio, mas a maioria é diagnosticada em função dos sintomas locais. Essas neoplasias são geralmente refratárias aos tratamentos atuais. Uma mulher com 51 anos de idade apresentou dor de cabeça de início súbito, paralisia de braço esquerdo e hipoestesia facial esquerda. A tomografia e a ressonância magnética revelaram um tumor pituitário invadindo os seios esfenoidal e cavernoso esquerdos. Os dados laboratoriais excluíram hipersecreção hormonal. A paciente foi submetida à cirurgia transesfenoidal, e os achados histológicos mostraram uma neoplasia com Ki-67 estimado em 75%. As imagens excluíram tanto um tumor oculto primário quanto disseminação sistêmica ou do sistema nervoso central. Três semanas após a cirurgia, a condição neurológica apresentou piora com início de ataxia, ptose bilateral, oftalmoplegia e aumento do tamanho da lesão, levando à intervenção cirúrgica por craniotomia, seguida por apenas algumas sessões de radioterapia devido à progressão grave da doença. A paciente veio a óbito depois de quase dois meses das manifestações iniciais. O caso ilustra a agressividade de algumas lesões pituitárias, a eficácia limitada das modalidades atuais de tratamento, como a cirurgia ou a radioterapia, e as limitações da classificação atual de tumores pituitários. Até onde sabemos, esse caso corresponde a uma das neoplasias pituitárias mais agressivas descritas até hoje, com um nível muito alto de Ki-67 (75%) e sobrevida curta (2 meses). O nível de Ki-67 pode ser de valor prognóstico em tumores pituitários. A classificação da Organização Mundial da Saúde (OMS) para tumores pituitários deveria ser revisitada.

The PROP1 2-Base Pair Deletion Is a Common Cause of Combined Pituitary Hormone Deficiency

Combined pituitary hormone deficiency (CPHD) has an incidence of approximately 1 in 8000 births. Although the proportion of familial CPHD cases is unknown, about 10% have an affected first degree relative. We have recently reported three mutations in the PROP1 gene that cause CPHD in human subjects. We report here the frequency of one of these mutations, a 301–302delAG deletion in exon 2 of PROP1, in 10 independently ascertained CPHD kindreds and 21 sporadic cases of CPHD from 8 different countries. Our results show that 55% (11 of 20) of PROP1 alleles have the 301–302delAG deletion in familial CPHD cases. Interestingly, although only 12% (5 of 42) of the PROP1 alleles of our 21 sporadic cases were 301–302delAG, the frequency of this allele (in 20 of 21 of the sporadic subjects given TRH stimulation tests) was 50% (3 of 6) and 0% (0 of 34) in the CPHD cases with pituitary and hypothalamic defects, respectively. Using whole genome radiation hybrid analysis, we localized the PROP1 gene to the distal end of chromosome 5q and identified a tightly linked polymorphic marker, D5S408, which can be used in segregation studies. Analysis of this marker in affected subjects with the 301–302delAG deletion suggests that rather than being inherited from a common founder, the 301–302delAG may be a recurring mutation.