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1.
Ultrasound Obstet Gynecol ; 57(2): 305-313, 2021 02.
Artigo em Inglês | MEDLINE | ID: mdl-31765050

RESUMO

OBJECTIVE: Insufflation of the amniotic cavity with carbon dioxide (CO2 ) is used clinically to improve visibility during complex fetoscopic surgery. Insufflation with heated, humidified CO2 has recently been shown to reduce fetal hypercapnia and acidosis in sheep, compared with use of cold and dry CO2 , but the underlying mechanisms are unclear. The aim of this study was to investigate whether differences in placental CO2 and oxygen (O2 ) exchange during insufflation with heated and humidified vs cold and dry CO2 could explain these findings. METHODS: Thirteen fetal lambs at 105 days of gestation (term, 146 days) were exteriorized partially, via a midline laparotomy and hysterotomy, and instrumented with an umbilical artery catheter, an umbilical vein catheter and a common umbilical vein flow probe. Arterial and venous catheters and flow probes were also inserted into the maternal uterine circulation. Six ewes were insufflated with cold, dry CO2 (22°C; 0-5% humidity) and seven with heated, humidified CO2 (40°C; 95-100% humidity) at 15 mmHg for 180 min. Blood-flow recordings and paired arterial and venous blood gases were sampled from uterine and umbilical vessels. Rates of placental CO2 and O2 exchange were calculated. RESULTS: After 180 min of insufflation, fetal survival was 33% (2/6) using cold, dry CO2 and 71% (5/7) using heated, humidified CO2 . By 120 min, fetuses insufflated with heated, humidified CO2 had lower arterial CO2 levels and higher arterial pH compared to those insufflated with cold, dry gas. Insufflation decreased significantly placental gas exchange in both groups, as measured by rates of both (i) fetal CO2 clearance and O2 uptake and (ii) maternal O2 delivery and CO2 uptake from the fetal compartment. CONCLUSIONS: Lower arterial CO2 and higher pH levels in fetuses insufflated with heated and humidified, compared to cold and dry, CO2 could not be explained by differences in placental gas exchange. Instead, heated and humidified insufflation appeared to reduce fetal CO2 absorption from the uterus, supporting its use in preference to cold, dry CO2 . © 2019 International Society of Ultrasound in Obstetrics and Gynecology.


Assuntos
Dióxido de Carbono/administração & dosagem , Insuflação , Placenta/metabolismo , Animais , Gasometria , Dióxido de Carbono/metabolismo , Feminino , Modelos Animais , Gravidez , Ovinos
2.
Ultrasound Obstet Gynecol ; 54(4): 506-516, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31364206

RESUMO

OBJECTIVES: Infants with congenital diaphragmatic hernia (CDH) are predisposed to pulmonary hypertension after birth, owing to lung hypoplasia that impairs fetal pulmonary vascular development. Antenatal sildenafil treatment attenuates abnormal pulmonary vascular and alveolar development in rabbit and rodent CDH models, but whether this translates to functional improvements after birth remains unknown. We aimed to evaluate the effect of antenatal sildenafil on neonatal pulmonary hemodynamics and lung function in lambs with diaphragmatic hernia (DH). METHODS: DH was surgically induced at approximately 80 days' gestation in 16 lamb fetuses (term in lambs is approximately 147 days). From 105 days' gestation, ewes received either sildenafil (0.21 mg/kg/h intravenously) or saline infusion until delivery (n = 8 fetuses in each group). At approximately 138 days' gestation, all lambs were instrumented and then delivered via Cesarean section. The lambs were ventilated for 120 min with continuous recording of physiological (pulmonary and carotid artery blood flow and pressure; cerebral oxygenation) and ventilatory parameters, and regular assessment of arterial blood gas tensions. Only lambs that survived until delivery and with a confirmed diaphragmatic defect at postmortem examination were included in the analysis; these comprised six DH-sildenafil lambs and six DH-saline control lambs. RESULTS: Lung-to-body-weight ratio (0.016 ± 0.001 vs 0.013 ± 0.001; P = 0.06) and dynamic lung compliance (0.8 ± 0.2 vs 0.7 ± 0.2 mL/cmH2 O; P = 0.72) were similar in DH-sildenafil lambs and controls. Pulmonary vascular resistance decreased following lung aeration to a greater degree in DH-sildenafil lambs, and was 4-fold lower by 120 min after cord clamping than in controls (0.6 ± 0.1 vs 2.2 ± 0.6 mmHg/(mL/min); P = 0.002). Pulmonary arterial pressure was also lower (46 ± 2 vs 59 ± 2 mmHg; P = 0.048) and pulmonary blood flow higher (25 ± 3 vs 8 ± 2 mL/min/kg; P = 0.02) in DH-sildenafil than in DH-saline lambs at 120 min. Throughout the 120-min ventilation period, the partial pressure of arterial carbon dioxide tended to be lower in DH-sildenafil lambs than in controls (63 ± 8 vs 87 ± 8 mmHg; P = 0.057), and there was no significant difference in partial pressure of arterial oxygen between the two groups. CONCLUSIONS: Sustained maternal antenatal sildenafil infusion reduced pulmonary arterial pressure and increased pulmonary blood flow in DH lambs for the first 120 min after birth. These findings of improved pulmonary vascular function are consistent with improved pulmonary vascular structure seen in two previous animal models. The data support the rationale for a clinical trial investigating the effect of antenatal sildenafil in reducing the risk of neonatal pulmonary hypertension in infants with CDH. Copyright © 2019 ISUOG. Published by John Wiley & Sons Ltd.


Assuntos
Hemodinâmica/efeitos dos fármacos , Hérnias Diafragmáticas Congênitas/tratamento farmacológico , Pulmão/efeitos dos fármacos , Inibidores da Fosfodiesterase 5/farmacologia , Citrato de Sildenafila/farmacologia , Animais , Autopsia/métodos , Gasometria/métodos , Feminino , Terapias Fetais/métodos , Feto , Hérnias Diafragmáticas Congênitas/fisiopatologia , Pulmão/irrigação sanguínea , Pulmão/fisiopatologia , Modelos Animais , Inibidores da Fosfodiesterase 5/administração & dosagem , Inibidores da Fosfodiesterase 5/sangue , Gravidez , Cuidado Pré-Natal , Troca Gasosa Pulmonar/efeitos dos fármacos , Ovinos , Citrato de Sildenafila/administração & dosagem , Citrato de Sildenafila/sangue
3.
Ultrasound Obstet Gynecol ; 53(3): 340-347, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30461102

RESUMO

OBJECTIVE: Partial amniotic carbon dioxide (CO2 ) insufflation (PACI) is used to improve visualization and facilitate complex fetoscopic surgery. However, there are concerns about fetal hypercapnic acidosis and postoperative fetal membrane inflammation. We assessed whether using heated and humidified, rather than cold and dry, CO2 might reduce the impact of PACI on the fetus and fetal membranes in sheep. METHODS: Twelve fetal lambs of 105 days' gestational age (term = 145 days) were exteriorized partially, via a midline laparotomy and hysterotomy, and arterial catheters and flow probes were inserted surgically. The 10 surviving fetuses were returned to the uterus, which was then closed and insufflated with cold, dry (22 °C at 0-5% humidity, n = 5) or heated, humidified (40 °C at 100% humidity, n = 5) CO2 at 15 mmHg for 180 min. Fetal membranes were collected immediately after insufflation for histological analysis. Physiological data and membrane leukocyte counts, suggestive of membrane inflammation, were compared between the two groups. RESULTS: After 180 min of insufflation, fetal survival was 0% in the group which underwent PACI with cold, dry CO2 , and 60% (n = 3) in the group which received heated, humidified gas. While all insufflated fetuses became progressively hypercapnic (PaCO2  > 68 mmHg), this was considerably less pronounced in those in which heated, humidified gas was used: after 120 min of insufflation, compared with those receiving cold, dry gas (n = 3), fetuses undergoing heated, humidified PACI (n = 5) had lower arterial partial pressure of CO2 (mean ± standard error of the mean, 82.7 ± 9.1 mmHg for heated, humidified CO2 vs 170.5 ± 28.5 for cold, dry CO2 during PACI, P < 0.01), lower lactate levels (1.4 ± 0.4 vs 8.5 ± 0.9 mmol/L, P < 0.01) and higher pH (pH, 7.10 ± 0.04 vs 6.75 ± 0.04, P < 0.01). There was also a non-significant trend for fetal carotid artery pressure to be higher following PACI with heated, humidified compared with cold, dry CO2 (30.5 ± 1.3 vs 8.7 ± 5.5 mmHg, P = 0.22). Additionally, the median (interquartile range) number of leukocytes in the chorion was significantly lower in the group undergoing PACI with heated, humidified CO2 compared with the group receiving cold, dry CO2 (0.7 × 10-5 (0.5 × 10-5 ) vs 3.2 × 10-5 (1.8 × 10-5 ) cells per square micron, P = 0.02). CONCLUSIONS: PACI with cold, dry CO2 causes hypercapnia, acidosis, hypotension and fetal membrane inflammation in fetal sheep, raising potential concerns for its use in humans. It seems that using heated, humidified CO2 for insufflation partially mitigates these effects and this may be a suitable alternative for reducing the risk of fetal acid-base disturbances during, and fetal membrane inflammation following, complex fetoscopic surgery. © 2018 The Authors. Ultrasound in Obstetrics & Gynecology published by John Wiley & Sons Ltd on behalf of the International Society of Ultrasound in Obstetrics and Gynecology.


Assuntos
Insuflação , Meningomielocele , Animais , Dióxido de Carbono , Feminino , Fetoscopia , Humanos , Modelos Animais , Gravidez , Ovinos , Útero
4.
Reprod Suppl ; 60: 9-18, 2002.
Artigo em Inglês | MEDLINE | ID: mdl-12220169

RESUMO

It has long been established that there are major variations in both the immunogenicity and antigenicity of native zona pellucida (ZP) proteins. These differences appear to be more pronounced with respect to genetically engineered ZP proteins, which do not have native post-translational modifications (for example glycosylation and sulphation). As the number of animal species that are now included in population management programmes using native porcine zona pellucida (PZP) proteins expands, it is increasingly important to carry out studies to evaluate the immune response variations among different species as well as the individual variation within a species. In an attempt to compare these complex immune responses, we have evaluated antibodies from numerous species immunized with native, genetically engineered ZP and synthetic ZP peptides. Such an immunocontraceptive method could have great potential. These studies are critical not only for the development of predictable immune responses that result in permanent sterilization versus reversible contraceptive effects, but also for predicting which vaccinogens (native ZP protein versus genetically engineered ZP proteins) might have detrimental effects on animal and human populations.


Assuntos
Anticorpos/imunologia , Anticoncepção Imunológica/veterinária , Proteínas do Ovo/administração & dosagem , Glicoproteínas de Membrana/administração & dosagem , Receptores de Superfície Celular , Vacinas Anticoncepcionais/imunologia , Animais , Anticorpos/sangue , Cães , Desenho de Fármacos , Epitopos/imunologia , Feminino , Cobaias , Haplorrinos , Camundongos , Mimetismo Molecular , Controle da População , Coelhos , Proteínas Recombinantes/imunologia , Suínos , Vacinas Anticoncepcionais/efeitos adversos , Vacinas Sintéticas/imunologia , Glicoproteínas da Zona Pelúcida
5.
Biol Reprod ; 65(3): 951-60, 2001 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-11514363

RESUMO

A lactosaminoglycan-associated antigen is associated with a carbohydrate moiety of all three zona pellucida (ZP) glycoproteins of pig and rabbit but is absent in the mouse and rat. A monoclonal antibody (PS1) recognizing this determinant was obtained by immunizing mice with a porcine ZP glycoprotein isoform purified by two-dimensional polyacrylamide gel electrophoresis. Conditions known to remove O-linked or sialic acid carbohydrate moieties (alkaline reduction; O-glycanase or neuraminidase enzymatic cleavage) did not remove the carbohydrate epitope. However, treatment with endo-beta-glycosidase, endoglycosidase F, or combinations of neuraminidase plus beta-galactosidase, totally removed the determinant, indicating that it is associated with a poly-N-acetyllactosaminoglycan structure present on an N-linked oligosaccharide. Molecular morphology studies using immunofluorescence and confocal microscopy techniques demonstrate that the PS1 antigen is localized at the surface of the ZP. Confirmation of this localization was obtained through studies that show that this antibody will inhibit homologous sperm binding to the pig ZP. Additional analyses using modular contrast microscopy and immunocytochemistry demonstrate that this carbohydrate-associated antigen is localized in discrete layers throughout the ZP matrix. These studies are the first to demonstrate the presence of a lactosaminoglycan type carbohydrate moiety in all three ZP proteins using a monoclonal antibody that appears to be involved in sperm recognition and structural organization.


Assuntos
Carboidratos/análise , Proteínas do Ovo/química , Glicosídeo Hidrolases , Glicoproteínas de Membrana/química , Receptores de Superfície Celular , Animais , Anticorpos Monoclonais/farmacologia , Antígenos/análise , Antígenos/imunologia , Configuração de Carboidratos , Carboidratos/química , Proteínas do Ovo/imunologia , Proteínas do Ovo/isolamento & purificação , Eletroforese em Gel Bidimensional , Feminino , Glicosilação , Hexosaminidases/metabolismo , Concentração de Íons de Hidrogênio , Hidrólise , Immunoblotting , Masculino , Glicoproteínas de Membrana/imunologia , Glicoproteínas de Membrana/isolamento & purificação , Camundongos , Microscopia Confocal , Neuraminidase/metabolismo , Coelhos , Hidróxido de Sódio , Espermatozoides/metabolismo , Suínos , Zona Pelúcida/metabolismo , Glicoproteínas da Zona Pelúcida , beta-Galactosidase/metabolismo
6.
J Soc Gynecol Investig ; 8(1 Suppl Proceedings): S43-8, 2001.
Artigo em Inglês | MEDLINE | ID: mdl-11223372

RESUMO

The ovary does not have a distinct morphologic barrier between the immune system and the developing gametes. This is in contrast to the testis in which the junctional complexes between the Sertoli cells form the blood-testis barrier. Whereas there are numerous factors, including genetic ones, associated with ovarian dysfunction, the immune factors have frequently been implicated in ovarian dysfunction. Much of our knowledge used to evaluate the immune system of the ovary has come from studies on the expression of the zona pellucida (ZP) proteins during ovarian development. Initial studies by Dunbar and colleagues demonstrated that immunization of rabbits with porcine ZP proteins (but not rabbit ZP proteins) would result in the generation of antibodies that inhibit sperm binding to the ZP and interfere with normal ovarian follicular development. In contrast to the rabbit and primate models, immunization of mice or rats with porcine ZP proteins does not have an effect on fertility or ovarian function although immunization of certain strains of mice with mouse ZP peptides and immune activator systems has been shown to result in ovarian pathology. Whereas immune inflammatory reactions have been observed in the mouse models, no such immune reactions have been observed in rabbit, guinea pig, or nonhuman primate models. Subsequent observations in nonhuman primates have shown that immunization of primates with ZP proteins expressed from cDNAs coding for the mouse and rabbit ZP2 (the mouse homologue has 60% amino acid identity with human ZP2) or the mouse ZP3 (the mouse protein has 67% amino acid identity with human ZP3) causes ovarian dysgenesis. In contrast, immunization of primates with recombinant rabbit ZP1 protein (the mouse homologue has 39% amino acid identity with human ZP1) does not affect nonhuman primate ovarian function or follicular development but will elicit antibodies that inhibit sperm binding to the primate ZP. These studies have collectively provided important information concerning the immunologic status of the ovary and demonstrate the species variations in immune responses to different ovarian immunogens.


Assuntos
Imunidade , Ovário/imunologia , Receptores de Superfície Celular , Animais , Autoantígenos/imunologia , Proteínas do Ovo/imunologia , Feminino , Humanos , Glicoproteínas de Membrana/imunologia , Doenças Ovarianas/imunologia , Ovário/crescimento & desenvolvimento , Zona Pelúcida/imunologia , Glicoproteínas da Zona Pelúcida
7.
South Med J ; 94(12): 1235, 2001 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-11811867
8.
Cells Tissues Organs ; 166(2): 148-64, 2000.
Artigo em Inglês | MEDLINE | ID: mdl-10729725

RESUMO

The zona pellucida (ZP) is the extracellular matrix that plays important roles in sperm-egg interaction. The ZP is composed of three major glycoproteins that exhibit heterogeneity due to extensive post-translational modifications including glycosylation and sulfation. Because of these modifications the nomenclature of ZP proteins from different species based on electrophoretic mobilities has been confusing. As the cDNAs and genes encoding the different ZP proteins have been isolated and sequenced, it is now possible to relate these ZP proteins according to gene families. Using the mouse ZP nomenclature, the ZP proteins from different mammalian species can be classified into three protein families: ZP1, ZP2, and ZP3. Although some of the structural domains of the ZP proteins of different species are conserved within each family, they exhibit distinct biological properties. In the mouse it has been established that ZP3 is the primary sperm receptor while ZP2 has secondary sperm receptor properties. In the pig, however, ZP1 has been shown to have sperm receptor activity similar to that observed in the rabbit and nonhuman primates. It is of interest that the human ZP2 and ZP3 gene families are 60-70% conserved with respect to the mouse ZP amino acid sequence, while the mouse ZP1 is only 39% conserved with respect to human ZP1. Such differences in protein structure and glysosylation may explain the marked species differences in the biochemical, physicochemical and immunochemical properties of the ZP. Studies have now shown that the proteins of the ZP are expressed in a stage specific manner and that there is increasing evidence that ZP proteins are expressed by both granulosa cells and the oocyte and may play a role in granulosa cell differentiation.


Assuntos
Proteínas da Matriz Extracelular/fisiologia , Interações Espermatozoide-Óvulo , Zona Pelúcida/fisiologia , Animais , Feminino , Humanos , Masculino , Mamíferos , Camundongos , Coelhos , Zona Pelúcida/ultraestrutura
9.
Biol Reprod ; 61(6): 1373-80, 1999 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-10569978

RESUMO

Zona pellucida (ZP) glycoproteins contain numerous antigenic determinants including carbohydrate, protein, and conformational epitopes; and the immunogenicity of these complex glycoproteins varies in different mammalian hosts. Studies have now shown that antibodies from primates immunized with a cDNA-expressed recombinant rabbit ZP protein (the homologue of the human ZP1 [hZP1]) inhibit sperm binding to the ZP without altering ovarian function, unlike immunization with ZP3 and ZP2 protein families. The ZP1 protein or peptides derived from it (recombinant or synthetic) are therefore primary candidates for use in designing safe and reversible human and animal contraceptive vaccines. In order to define peptide epitope(s) that may be critical for eliciting an immune response sufficient to effect immunological contraception without causing any adverse effects on ovarian physiology, studies have been carried out to identify immunodominant B-cell epitopes of the ZP1 protein. The amino acid sequence of the hZP1 was used to design a set of 94 (15-mer) biotinylated peptides having an overlap of 9 amino acids. Using these peptides in a modified enzyme-linked immunoassay, antibodies in sera from rabbits or baboons immunized with native porcine ZP protein were screened for ZP1 peptide recognition. These studies demonstrate that there are a limited number of peptides recognized by primate antibodies but that the overlapping peptides sharing the sequence GPLTLELQI are recognized by both rabbit and baboon antibodies regardless of the adjuvant system used to induce the immune response. This peptide is 100% conserved in amino acid sequence between the human and pig, although the rabbit protein has two conserved amino acid substitutions (100% similar, 77% identical). Because this peptide is immunogenic as well as antigenic in primates, it could play a major role in the development of human contraceptive vaccines.


Assuntos
Linfócitos B/imunologia , Proteínas do Ovo/imunologia , Epitopos/imunologia , Glicoproteínas de Membrana/imunologia , Receptores de Superfície Celular , Sequência de Aminoácidos , Animais , Biotinilação , Proteínas do Ovo/análise , Proteínas do Ovo/química , Eletroforese em Gel Bidimensional , Epitopos/análise , Epitopos/química , Feminino , Humanos , Glicoproteínas de Membrana/análise , Glicoproteínas de Membrana/química , Dados de Sequência Molecular , Papio/imunologia , Peptídeos/química , Peptídeos/imunologia , Coelhos , Especificidade da Espécie , Glicoproteínas da Zona Pelúcida
11.
FEBS Lett ; 413(2): 215-25, 1997 Aug 18.
Artigo em Inglês | MEDLINE | ID: mdl-9280285

RESUMO

The primary structure of two forms of gonadotropin releasing hormone (GnRH) from tunicate (Chelyosoma productum) have been determined based on mass spectrometric and chemical sequence analyses. The peptides, tunicate GnRH-I and -II, contain features unprecedented in vertebrate GnRH. Tunicate GnRH-I contains a putative salt bridge between Asp5 and Lys8. A GnRH analog containing a lactam bridge between Asp5 and Lys8 was found to increase release of estradiol compared with that of the native tunicate GnRH-I and -II. Tunicate GnRH-II contains a cysteine residue and was isolated as a dimeric peptide. These motifs suggest that the conformation plays an important role in receptor activation.


Assuntos
Hormônio Liberador de Gonadotropina/química , Receptores LHRH/metabolismo , Urocordados/química , Sequência de Aminoácidos , Animais , Dimerização , Estradiol/metabolismo , Hormônio Liberador de Gonadotropina/análogos & derivados , Hormônio Liberador de Gonadotropina/farmacologia , Fragmentos de Peptídeos/química , Conformação Proteica , Análise de Sequência , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Urocordados/metabolismo
12.
Anticancer Res ; 17(2A): 901-6, 1997.
Artigo em Inglês | MEDLINE | ID: mdl-9137425

RESUMO

A monoclonal antibody developed against a meiotically expressed porcine oocyte carbohydrate antigen has been shown to recognize an antigen in ovarian surface epithelial cells. Immunohistochemical studies of ovaries demonstrated that this antigen is present in the ovarian surface epithelia (OSE) of numerous mammalian species, including the non-human primate and the human (1). Although most of the ovarian surface epithelial cells are lost during aging in the human, a few cells may remain in ovarian crypts. In view of theories that most ovarian carcinomas are derived from the OSE cells in aging women, the PS1 antibody has been used to evaluate ovarian tumors using immunocytochemistry to detect the PS1 antigen in paraffin embedded pathology tissues. The present study found that the PS1 antigen is abundant in a number of malignant ovarian tumors, but is not expressed in a non-malignant Brenner's (ovarian) tumor or granulosa cell tumors. This antibody therefore appears to have great potential for the histopathological and immunochemical analysis of ovarian tumors.


Assuntos
Antígenos Glicosídicos Associados a Tumores/análise , Neoplasias Ovarianas/imunologia , Animais , Anticorpos Monoclonais/imunologia , Feminino , Humanos , Imuno-Histoquímica , Meiose , Camundongos
13.
Anticancer Res ; 17(2A): 907-11, 1997.
Artigo em Inglês | MEDLINE | ID: mdl-9137426

RESUMO

A monoclonal antibody developed against a meiotically expressed porcine oocyte carbohydrate antigen has been shown to recognize an antigen in ovarian surface epithelial cells (OSE) of numerous mammalian species, including the non-human primate and the human (1). Although most of the ovarian surface epithelial cells are lost during aging in the human, a few cells may remain in ovarian crypts. Because the majority of ovarian carcinomas are thought to be derived from the OSE cells in aging women the PS1 antibody has been used to evaluate ovarian tumors. The secretory origin of this carbohydrate antigen in meiotic cells prompted further analyses of peritoneal fluid collected from gynecological surgery patients including those diagnosed with ovarian cancer. The present study demonstrates that ovarian tumor proteins separated on SDS PAGE include an antigen having a heterogeneous molecular weight (> 100 kDa) typical of glycosylated proteins. Additional studies show that peritoneal fluid from 19 patients not having cancer contain PS1 associated glycoproteins. However, of 14 cancer patients, only one had detectable levels of the carbohydrate antigen. These observations suggest that either the secretion of this glycoprotein is altered in ovarian carcinoma or that glycosidases or other proteolytic enzymes are involved in the degradation of these glycoproteins.


Assuntos
Antígenos Glicosídicos Associados a Tumores/análise , Líquido Ascítico/química , Proteínas de Neoplasias/análise , Neoplasias Ovarianas/química , Animais , Feminino , Humanos , Meiose , Camundongos
14.
Proc Natl Acad Sci U S A ; 93(19): 10461-4, 1996 Sep 17.
Artigo em Inglês | MEDLINE | ID: mdl-8816823

RESUMO

The neuropeptide gonadotropin-releasing hormone (GnRH) is the major regulator of reproduction in vertebrates. Our goal was to determine whether GnRH could be isolated and identified by primary structure in a protochordate and to examine its location by immunocytochemistry. The primary structure of two novel decapeptides from the tunicate Chelyosoma productum (class Ascidiacea) was determined. Both show significant identity with vertebrate GnRH. Tunicate GnRH-I (pGlu-His-Trp-Ser-Asp-Tyr-Phe-Lys-Pro-Gly-NH2) has 60% of its residues conserved, compared with mammalian GnRH, whereas tunicate GnRH-II (pGlu-His-Trp-Ser-Leu-Cys-His-Ala-Pro-Gly-NH2) is unusual in that it was isolated as a disulfide-linked dimer. Numerous immunoreactive GnRH neurons lie within blood sinuses close to the gonoducts and gonads in both juveniles and adults, implying that the neuropeptide is released into the bloodstream. It is suggested that in ancestral chordates, before the evolution of the pituitary, the hormone was released into the bloodstream and acted directly on the gonads.


Assuntos
Evolução Biológica , Hormônio Liberador de Gonadotropina/química , Sistema Nervoso/química , Urocordados/fisiologia , Sequência de Aminoácidos , Animais , Galinhas , Dissulfetos , Peixes , Gânglios dos Invertebrados/química , Hormônio Liberador de Gonadotropina/genética , Hormônio Liberador de Gonadotropina/isolamento & purificação , Mamíferos , Dados de Sequência Molecular , Especificidade de Órgãos , Homologia de Sequência de Aminoácidos , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Urocordados/genética
15.
Aust Dent J ; 41(3): 188-92, 1996 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-8768644

RESUMO

A low power Ga-As pulse laser was used to stimulate cultured human embryonic fibroblast cells. Energy fluencies varied from 0-1 J/cm2 over a period of 1-4 days. Fibroblast procollagen production was monitored by the synthesis of [3H] hydroxyproline, and DNA replication was assessed by [3H] thymidine incorporation. Following laser treatment, controlled pepsin digestion measured the increase in cell biostimulation. Maximum increase in collagen production and cell biostimulation occurred after 4 episodes of laser treatment at 24-hour intervals. Laser doses between 0.099 and 0.522 J/cm2 had the most significant stimulatory effects on fibroblast function. Clinical efficacy of the low power Ga-As pulse laser may be related to enhanced connective tissue repair.


Assuntos
Colágeno/metabolismo , Fibroblastos/metabolismo , Lasers , Arsênio , Células Cultivadas , Colágeno/efeitos da radiação , Tecido Conjuntivo/metabolismo , Tecido Conjuntivo/efeitos da radiação , Replicação do DNA/efeitos da radiação , Embrião de Mamíferos , Fibroblastos/efeitos da radiação , Gálio , Humanos , Hidroxiprolina/biossíntese , Hidroxiprolina/efeitos da radiação , Pepsina A , Pró-Colágeno/biossíntese , Pró-Colágeno/metabolismo , Pró-Colágeno/efeitos da radiação , Timidina/metabolismo , Timidina/efeitos da radiação , Fatores de Tempo , Trítio
16.
Mol Reprod Dev ; 43(4): 519-29, 1996 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-9052944

RESUMO

A cDNA encoding the rabbit 55 kDa ZP protein was expressed using a baculovirus expression system and was evaluated for its ability to elicit antibodies which may interfere with sperm-ZP interaction. The expressed glycosylated protein, BV55, was purified by wheat germ agglutinin lectin affinity chromatography. Antisera made in guinea pigs immunized with BV55 (GP-alpha-BV55) is specific for the 55 kDa rabbit ZP protein. Indirect immunofluorescence studies indicate that GP-alpha-BV55 localizes to a filamentous meshwork on the surface of the ZP of isolated rabbit eggs. Immunohistochemical analysis of rabbit ovaries demonstrated that this antigen is localized within the ZP of primary and more advanced stage ovarian follicles but is not detected in primordial follicles. In addition, the 55 kDa antigen was detected in the granulosa cells of secondary stage follicles but not in the oocyte. GP-alpha-BV55 effectively blocked the binding of rabbit sperm to rabbit eggs in vitro. However, Fab fragments generated from GP-alpha-BV55 failed to block sperm binding, suggesting that the inhibitory effect of GP-alpha-BV55 was due to stearic hindrance rather than specific blocking of a sperm receptor site. Although the Fab fragment did not inhibit sperm binding, additional studies demonstrated that biotinylated BV55 protein bound to rabbit sperm in the acrosomal region in a manner consistent with ligand activity in the sperm-ZP interaction, and that BV55 bound to rabbit sperm in a dose-dependent manner. These studies therefore demonstrate that antibodies against recombinant ZP proteins recognize the native intact ZP and inhibit sperm-ZP interaction. They also provide evidence that the rabbit 55 kDa ZP protein, which is the homolog of the pig ZP3 alpha sperm receptor protein, has sperm receptor activity.


Assuntos
Proteínas do Ovo/metabolismo , Glicoproteínas de Membrana/metabolismo , Receptores de Superfície Celular , Zona Pelúcida/metabolismo , Animais , Anticorpos , Linhagem Celular , Proteínas do Ovo/química , Proteínas do Ovo/genética , Ensaio de Imunoadsorção Enzimática , Feminino , Técnica Indireta de Fluorescência para Anticorpo , Vetores Genéticos , Cobaias , Immunoblotting , Masculino , Glicoproteínas de Membrana/química , Glicoproteínas de Membrana/genética , Nucleopoliedrovírus/genética , Espermatozoides/metabolismo , Spodoptera/citologia , Glicoproteínas da Zona Pelúcida
19.
Int J Dermatol ; 33(9): 628-31, 1994 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-8002157

RESUMO

BACKGROUND: Chronic myelomonocytic leukemia has been associated with various nonspecific cutaneous manifestations. Rarely has the leukemia been reported to directly affect the skin. METHODS: This case documents the progression of a patient who ultimately developed chronic myelomonocytic leukemia, by clinical examination, hematologic parameters, dermatopathology, and bone marrow pathology. RESULTS: The skin showed nonspecific cutaneous involvement, progressing to specific leukemic lesions parallel with increasing systemic and hematologic involvement. CONCLUSIONS: Chronic myelomonocytic leukemia can manifest with lesions of leukemia cutis. The possibility of nonspecific cutaneous involvement in the preleukemic phase exists.


Assuntos
Leucemia Mielomonocítica Crônica/complicações , Leucemia Mielomonocítica Crônica/diagnóstico , Dermatopatias/diagnóstico , Dermatopatias/etiologia , Biópsia , Contagem de Células Sanguíneas , Evolução Fatal , Testes Hematológicos , Humanos , Leucemia Mielomonocítica Crônica/terapia , Masculino , Pessoa de Meia-Idade , Dermatopatias/fisiopatologia , Dermatopatias/terapia
20.
J Reprod Fertil ; 101(2): 295-303, 1994 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-7523667

RESUMO

The effectiveness of zona pellucida antigens in immunizing white-tailed deer to reduce fertility was evaluated by analysing the constituent deer zona pellucida proteins and their immunogenicity. Does were immunized with porcine zona pellucida antigens. The antibodies were characterized using immunohistochemical and immunoblot analysis, in which zona pellucida proteins were separated by one-dimensional and two-dimensional PAGE. Deer anti-porcine zona pellucida antibodies were found to recognize all the major proteins of the porcine zona pellucida. These antibodies also recognized several proteins of deer zona pellucida, indicating that it is possible to break immune tolerance in the deer using such a protocol. The antibodies were also found to recognize peptides of 55 and 75 kDa that were produced by expressing cDNA clones containing antigens of major glycoproteins of rabbit zona pellucida. Furthermore, antibodies against rabbit zonae pellucidae recognized antigens in zonae of paraffin-embedded deer ovaries. Taken together, these experiments demonstrate the crossreactive nature of a number of zona pellucida epitopes found in deer and in several other species. They also illustrate the immunogenicity possible in such an immunization protocol, and provide valuable probes for the investigation of follicular development in this and other species.


Assuntos
Anticorpos/sangue , Antígenos/administração & dosagem , Cervos/imunologia , Proteínas do Ovo/imunologia , Fertilidade/imunologia , Zona Pelúcida/imunologia , Animais , Reações Cruzadas , Eletroforese em Gel Bidimensional , Eletroforese em Gel de Poliacrilamida , Epitopos/imunologia , Feminino , Imunização , Coelhos , Proteínas Recombinantes/imunologia , Suínos
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