Transverse myelitis in patients with antiphospholipid antibodies--the importance of early diagnosis and treatment.

Transverse myelitis (TM) is a rare manifestation of systemic lupus erythematosus (SLE) and the antiphospholipid syndrome (APS). No uniform therapeutic protocol exists for its treatment, and the prognosis is usually poor. Here we describe four patients having TM associated with antiphospholipid antibodies. Treatment measures and delay in diagnosis between symptom onset and the initiation of treatment varied between patients, but the earlier the diagnosis and the more aggressive the treatment the better was the patient's outcome. Based on these cases and on a literature review we suggest that early aggressive treatment (usually with pulses of methylprednisolone and cyclophosphamide) might improve the prognosis of patients with TM associated with antiphospholipid antibodies.

Seasonal fluctuations in pituitary levels of the three forms of gonadotropin-releasing hormone in striped bass, Morone saxatilis (Teleostei), during juvenile and pubertal development.

In most vertebrates, the development of a mature gonadotropin-releasing hormone (GnRH) secretory system is pivotal for the onset of puberty. The role of the three native GnRH forms, seabream (sb) GnRH, chicken (c) GnRH-II and salmon GnRH, in striped bass puberty remains elusive. This study examined the changes in pituitary GnRH levels throughout juvenile and pubertal development, a period encompassing 3 to 4 years. The levels of the two most abundant forms in the pituitary, sbGnRH and cGnRH-II (10:1), increased during the Fall and peaked prior to (cGnRH-II) or during (sbGnRH) the natural breeding season in March to May. In most cases, sbGnRH and cGnRH-II levels of maturing fish correlated to changes in oocyte diameter, gonadosomatic index and LH pituitary content. Interestingly, pituitaries of immature and maturing 2- and 3-year-old males and females contained similar amounts of all three GnRH forms. Additionally, pituitary sbGnRH and cGnRH-II levels in juvenile fish were relatively high and GnRH profiles showed a clear seasonality, similar to those of older, mature fish. These findings suggest a role for both sbGnRH and cGnRH-II in the regulation of gonadal development and indicate that, unlike some mammalian species, the timing of puberty in striped bass is not limited by a low activity of the GnRH system.

Early maturity in the male striped bass, Morone saxatilis: follicle-stimulating hormone and luteinizing hormone gene expression and their regulation by gonadotropin-releasing hormone analogue and testosterone.

Striped bass are seasonal breeding fish, spawning once a year during the spring. All 3-yr-old males are sexually mature; however, 60-64% of the fish mature earlier as 1- or 2-yr-old animals. The endocrine basis underlying early maturity in 2-yr-old males was studied at the molecular level by monitoring changes in pituitary beta FSH and beta LH mRNA levels by ribonuclease protection assay, and correlating these changes to stages of testicular development. In maturing males, the mRNA levels of beta FSH were elevated during early spermatogenesis, whereas beta LH mRNA levels peaked during spermiation. The appearance of spermatozoa in the testis was associated with a decrease in beta FSH mRNA and a rise in beta LH mRNA abundance. Immature males had lower levels of beta LH mRNA than maturing males, but there were no differences in beta FSH mRNA levels between immature and maturing males. The regulation of gonadotropin gene expression in 2-yr-old males was studied by the chronic administration of GnRH analogue (GnRHa) and testosterone (T), with or without pimozide (P) supplementation. In immature males, the combination of T and GnRHa stimulated a three- to fivefold increase in beta FSH and beta LH mRNA levels, but the same treatment had no effect on gonadotropin gene expression in maturing males. In addition, the coadministration of P to immature males suppressed the stimulatory effect of GnRHa and T on beta FSH and beta LH mRNA levels, suggesting that dopamine may have a novel role in regulating gonadotropin gene expression.

Characterization of a pituitary GnRH-receptor from a perciform fish, Morone saxatilis: functional expression in a fish cell line.

Gonadotropin-releasing hormones (GnRHs) bind to the specific receptor present on the gonadotrophs to activate the synthesis and release of gonadotropins (follicle stimulating hormone or FSH and luteinizing hormone or LH), which in turn control gonadal maturation, gametogenesis and gamete release. Perciform species have three endogenous GnRHs. The main objective of this study was to characterize the gonadotropin-releasing hormone receptor (GnRH-R) present in the pituitary of a perciform species, striped bass (Morone saxatilis) and demonstrate how it interacts with its potential ligand. In this study, a cDNA for GnRH-R from the pituitaries of striped bass was cloned. The cloned cDNA has an open reading frame (ORF) that codes for a 419 amino acids peptide. Like other G-protein coupled receptors including the non-mammalian GnRH-Rs, the peptide has seven putative transmembrane domains and a C-terminal tail. Comparative analysis of the amino acid sequence of striped bass (stb) GnRH-R shows 38-87% similarity with the known GnRH-Rs. A Northern blot analysis revealed a single GnRH-R transcript in the pituitary; however, its expression in various extrapituitary tissues was demonstrated by a reverse-transcription-PCR (RT-PCR). Functionally, upon induction by endogenous forms of GnRHs (seabream, chicken II and salmon GnRHs) and a mammalian GnRH-agonist, the recombinant stbGnRH-R mediated a reporter gene (luciferase) activity in a fish cell line (CHSE-214). A real-time relative quantitation method established that significantly higher (P<0.05) levels of stbGnRH-R mRNA were present in the pituitaries of striped bass with advanced stages of ovarian development, compared to the pituitaries of fish with less developed ovaries.

Gonadal development and plasma steroid levels during pubertal development in captive-reared striped bass, Morone saxatilis.

Puberty is the period during which full sexual development occurs and the capacity to reproduce is acquired. Despite its importance, our understanding of the endocrine regulation of puberty in lower vertebrates is still limited. The objective of the present study was to describe the changes in gonadal development and plasma steroid levels in a relatively late maturing species, the striped bass, during the first four years of life. In about 65% of the females, puberty was initiated during the third year. Although gonadosomatic index (GSI) and oocyte diameter increased during this year, this first cycle was characterized by a heterogeneous population of developing oocytes, a relatively low mean maximum oocyte diameter, and an absence of yolk granules in the oocytes. Plasma 17beta-estradiol (E(2)) levels were low in all three-year-old fish, suggesting that an insufficient stimulation of vitellogenin production by E(2) may underlie the lack of vitellogenin incorporation into developing oocytes. All monitored parameters increased during the fourth year, but were still below the values attained by older females. In about 60% of the males, puberty was initiated during the first year and all males were mature by the third year. During the first two years, several immature males initiated spermatogenesis without reaching full maturity. In mature males, mean GSI, plasma testosterone, and 11-ketotestosterone levels increased simultaneously, reaching higher values each subsequent year. Our results indicate that, similar to the situation in mammals, more than one reproductive cycle is required in striped bass before complete adulthood is reached.

Ontogeny of follicle-stimulating hormone and luteinizing hormone gene expression during pubertal development in the female striped bass, Morone saxatilis (Teleostei).

Pubertal development in teleost fish is characterized by gonadal growth that is directly stimulated by the pituitary gonadotropins, FSH and LH. We used a quantitative ribonuclease protection assay to provide, for the first time, the developmental profiles of the alpha-, betaFSH-, and betaLH-subunit gene expression in a seasonal breeding fish, the female striped bass (3-yr study, n = 207). Two-year-old females were sexually immature, although a transient rise in all gonadotropin subunit mRNAs was measured in the pituitary. Pubertal ovarian development occurred in 65% of 3-yr-old females, characterized by the appearance of lipid droplets within the oocytes. This reproductive phase, termed pubertal development, was associated with a 34-fold increase in the mRNA levels of betaFSH and a rise in the pituitary concentration of LH. The first sexual maturation took place in 4-yr-old females and coincided with a 218-fold increase in the mRNA levels of betaFSH. During this time period, the mRNA levels of the alpha and betaLH subunits increased by 11- and 8-fold, respectively. At the final stages of vitellogenic growth, mRNA levels of betaFSH declined to basal levels, whereas the mRNA levels of the alpha and betaLH subunits remained elevated. Throughout the study, pituitary LH concentration was positively correlated to the mRNA levels of betaLH, but plasma levels of LH remained low and unchanged (0.4-0.8 ng/ml) despite increasing levels of pituitary LH concentration, suggesting a regulated secretion pathway. Taken together, the data show that the profiles of betaFSH and betaLH mRNAs appear to follow an annual rhythm that is associated with developmental events in the growing oocytes. In particular, increasing levels of betaFSH mRNA appear to underlie the first sexual maturity in the female striped bass.

Effects of long-term testosterone, gonadotropin-releasing hormone agonist, and pimozide treatments on gonadotropin II levels and ovarian development in juvenile female striped bass (Morone saxatilis).

The ability of the juvenile female reproductive axis to respond to hormonal stimulation was investigated in a Perciform fish, the striped bass (Morone saxatilis) using various combinations of testosterone (T), GnRH agonist (GnRHa), and pimozide. A long-term treatment with T alone, or T in combination with GnRHa, increased pituitary gonadotropin II (GtH II) levels 2- and 3-fold, respectively, suggesting that T and GnRHa each stimulate GtH II accumulation. Release of the accumulated GtH II could be induced only by high doses of GnRHa in combination with T, indicating that GtH II synthesis and release require different levels of GnRH stimulation. The addition of the dopamine antagonist pimozide did not affect pituitary and plasma GtH II levels but, in response to an additional acute GnRHa challenge, inhibited the release of GtH II. Although ovarian development was slightly stimulated by a combined T and GnRHa treatment, vitellogenesis was generally not initiated. The present study demonstrated that the juvenile striped bass pituitary is responsive to hormonal stimulation, resulting in elevated levels of GtH II in the pituitary and plasma. However, increased plasma levels of GtH II did not result in precocious puberty, suggesting that additional factors are required for the initiation of ovarian development in this teleost.

Gonadotropin-I and -II subunit gene expression of male striped bass (Morone saxatilis) after gonadotropin-releasing hormone analogue injection: quantitation using an optimized ribonuclease protection assay.

In fish, both gonadotropin (GtH)-I and -II are involved in the spermatogenic process, but the differential regulation of these hormones by GnRH is still poorly understood. To gain further insight into the GnRH regulation of GtH-I and -II gene expression in the male striped bass, we have developed and optimized a ribonuclease protection assay for the simultaneous measurement of all GtH subunit mRNAs in a single pituitary gland. The RNA extraction protocol enables the determination of GtH protein content in the same sample, thus enhancing the power of the method. Maturing striped bass males were injected intramuscularly with [D-Ala6,Pro9Net]-LHRH (GnRHa) and sampled at 6 and 24 h postinjection. The mRNA levels of the alpha subunit and GtH-IIbeta increased after 6 h (4- and 6-fold, respectively), while the GtH-Ibeta mRNA levels increased only 2-fold after 24 h. Interestingly, GnRHa stimulation caused a significant increase in beta-actin mRNA levels. GnRHa treatment also resulted in a 2-fold decrease in pituitary GtH-II content, associated with a dramatic increase of plasma GtH-II levels from undetectable levels (< 0.2 ng/ml) to 13+/-2 ng/ml after 6 h. These results demonstrate that both GtH-Ibeta and -Ilbeta are expressed during striped bass spermatogenesis and that the two genes are subjected to differential regulation by GnRHa.

Gonadotropins beta-GtHI and beta-GtHII from the gilthead seabream, Sparus aurata.

Full-length cDNA sequences encoding the beta-subunits of the gonadotrophins GtHI and GtHII were isolated, cloned, and sequenced from a single gilthead seabream (Sparus aurata) pituitary using RACE PCR. Beta-GtHI and beta-GtHII degenerate PCR primers were designed according to regions of high amino acid sequence homology between the chum salmon and the bonito beta-GtHI or beta-GtHII. DNA sequence analysis of the cloned PCR products confirmed the presence of the predicted complete coding region as well as 5' and 3' untranslated sequences. The deduced amino acid sequences of beta-GtHI and beta-GtHII from the gilthead seabream were compared to GtH sequences from a number of teleosts. Seabream beta-GtHI shows the highest homology to the bonito beta-GtHI (76%) and the striped bass (74%), while seabream beta-GtHII is 98% homologous to the yellow fin porgy, 87% homologous to the bonito, and 93% homologous to the striped bass beta-GtHII. Northern blot analysis showed the transcripts of the gilthead seabream beta-GtHI and beta-GtHII to be about 700 and 740 bases, respectively. During the spawning season, beta-GtHI is expressed at higher levels in males than in females while beta-GtHII is expressed at similar levels in both sexes.

Molecular cloning and sequence analysis of striped bass (Morone saxatilis) gonadotrophin-I and -II subunits.

Two types of cDNA, each encoding a different beta-subunit of striped bass (Morone saxatilis, Teleostei) gonadotrophins (GTH-I beta and GTH-II beta), as well as the glycoprotein alpha-subunit, were cloned by screening a striped bass pituitary cDNA library. The probes used for screening the library were cloned cDNA fragments, generated by PCR amplification of reverse-transcribed mRNA obtained from two pituitaries. The nucleotide sequences of the alpha-subunit, GTH-I beta and GTH-II beta are 626, 524 and 580 bases long, encoding peptides of 117, 120 and 147 amino acids respectively. Striped bass GTH-I beta and GTH-II beta share a sequence identity of 48% at the nucleic acid level, and 30% at the amino acid level. A cluster analysis of vertebrate pituitary glycoprotein beta-subunits suggests that teleost GTH-II beta is more closely related to tetrapod LH than to FSH. Administration of gonadotrophin-releasing hormone analogue ([D-Ala6,Pro9Net]-LHRH) to juvenile striped resulted in ten-, two- and fivefold increases in the expression of the alpha-subunit, GTH-I beta and GTH-II beta respectively. These results suggest that each of the GTH subunits is differentially regulated, and further corroborate the functional duality of teleost gonadotrophins.