RESUMO
BACKGROUND: The biological potency of allergens can be measured by provoking mediator release from effector cells. As established immunochemical methods in allergen standardization only determine inhibition potency or major allergen content, routine tests for biological potency may enhance standardization and batch control of allergen products. OBJECTIVE: The general performance and application potential of biological in vitro assays in batch control and standardization of allergens and as a tool for verifying activity and stability of allergen standards were analysed. METHODS: Allergen extracts of five clinically relevant allergens from three to five different manufacturers were investigated. A CAP-IgE-inhibition assay was compared with mediator release assay (MRA)s based on murine or human basophils. Rat basophilic leukaemia (RBL) cells were passively sensitized with pooled murine allergen-specific IgE-containing sera. Humanized RBL cells and human-stripped basophils were sensitized with pooled patient's sera, which were also used for the CAP-IgE-inhibition assay. Allergen specificity of the sera was determined by immunoblotting. RESULTS: A good batch-to-batch consistency was found with each assay among all manufacturers and allergens tested. Between different manufacturers, the products showed differences in activity and the various assays indicated an almost identical ranking. However, the biological assays revealed qualitative differences of biological activity or composition of allergen preparations undetectable by IgE-inhibition assay. CONCLUSIONS: MRAs provide refined information on allergen activity, either confirming the results of IgE-inhibition assay, or indicating differences requiring further investigation, and represent a highly sensitive novel tool in allergen standardization. By using permanently cultivated cell lines, repeated venepuncture to obtain human basophils is avoided. As in the RBL assay, the coefficient of variation for the release values were below 15% and for the ED50 below 25%, the assay is suitable to determine differences that are relevant for batch control purposes.
Assuntos
Alérgenos/análise , Imunoglobulina E/imunologia , Alérgenos/imunologia , Animais , Especificidade de Anticorpos , Betula , Reações Cruzadas , Relação Dose-Resposta Imunológica , Humanos , Immunoblotting , Indicadores e Reagentes , Camundongos , Poaceae , Ratos , Kit de Reagentes para Diagnóstico , Padrões de Referência , Sensibilidade e EspecificidadeRESUMO
The mGluR5 antagonist 2-methyl-6-(phenylethynyl) pyridine (MPEP) produces anxiolytic or antidepressant effects in several rodent models through incompletely described mechanisms. Anxiolytics and antidepressants share several neuroendocrine features, including acute activation of the hypothalamic-pituitary-adrenal (HPA)-axis, desensitization of neuroendocrine responses with repeated dosing, and desensitization of the HPA axis to 5-HT1A agonist stimulation. We characterized these neuroendocrine parameters in rats treated systemically with MPEP and compared them to those induced by the anxiolytic buspirone. Acutely, MPEP dose-dependently (0.1-10 mg/kg i.p.) increased plasma corticosterone concentrations. These responses were blocked by 50% with the 5-HT1A antagonist WAY100635. The corticosterone responses to both 3 mg/kg MPEP and buspirone were decreased by 80% after 5 days of twice-daily injections. Repeated injection with MPEP decreased HPA-axis sensitivity to buspirone challenge by 75%. This desensitization was not associated with changes in mGluR5 or 5-HT1A receptor binding properties, expression of G-protein subunits coupled to these receptors, or in 5-HT-stimulated binding of [(3)H]-GTPgammaS to membranes. We conclude that MPEP acutely disinhibits the HPA axis, in part through uncharacterized changes in serotonergic signaling. Desensitization of 5-HT1A responses after repeated MPEP administration may indicate that, like other anxiolytics and antidepressants, plasticity in 5-HT signal transduction pathways has occurred.
Assuntos
Antagonistas de Aminoácidos Excitatórios/farmacologia , Sistema Hipotálamo-Hipofisário/efeitos dos fármacos , Sistema Hipófise-Suprarrenal/efeitos dos fármacos , Receptores de Glutamato Metabotrópico/antagonistas & inibidores , Serotonina/farmacologia , Animais , Relação Dose-Resposta a Droga , Sistema Hipotálamo-Hipofisário/metabolismo , Masculino , Sistema Hipófise-Suprarrenal/metabolismo , Piridinas/farmacologia , Ratos , Ratos Sprague-Dawley , Receptor de Glutamato Metabotrópico 5 , Receptores de Glutamato Metabotrópico/metabolismo , Receptores de Serotonina/metabolismo , Receptores 5-HT1 de Serotonina , Serotonina/metabolismo , Agonistas do Receptor de Serotonina/farmacologiaRESUMO
Recent studies in mice have clearly demonstrated that eliminating Apo E alters the rate, character and distribution of A beta deposits. In the present study, we asked whether elevating the levels of Apo E can, in a dominant fashion, influence amyloid deposition. We expressed human (Hu) Apo E4 via the mouse prion protein promoter, resulting in high expression in both astrocytes and neurons; only astrocytes efficiently secreted Hu Apo E4 (at least 5-fold more than endogenous). Mice hyper-expressing Hu Apo E4 developed normally and lived normal lifespans. The co-expression of Hu Apo E4 with a mutant amyloid precursor protein (APP) (Mo/Hu APPswe) or mutant APP and mutant presenilin (PS1dE9) did not lead to proportional changes in the age of appearance, relative burden, character or distribution of A beta deposits. We suggest that these data are best explained by proposing that the mechanisms by which Apo E influences A beta deposition involves an aspect of its normal function that is not augmented by hyper-expression.
Assuntos
Apolipoproteínas E/metabolismo , Astrócitos/metabolismo , Neurônios/metabolismo , Peptídeos beta-Amiloides/metabolismo , Precursor de Proteína beta-Amiloide/genética , Precursor de Proteína beta-Amiloide/metabolismo , Animais , Apolipoproteína E4 , Apolipoproteínas E/genética , Astrócitos/citologia , Encéfalo/metabolismo , Células Cultivadas , Regulação da Expressão Gênica , Humanos , Immunoblotting , Camundongos , Camundongos Transgênicos , Mutação , Neurônios/citologia , Fatores de TempoRESUMO
The amyloid precursor protein undergoes proteolysis at several sites to yield a number of functionally relevant peptides, including beta-amyloid and the soluble amyloid precursor protein derivatives alpha-soluble amyloid precursor protein and beta-soluble amyloid precursor protein. beta-Amyloid is the primary constituent of senile plaques associated with Alzheimer's disease, while a-soluble amyloid precursor protein promotes synaptogenesis and plays a role in neuroprotective processes. We tested for age-related alterations in these amyloid precursor protein proteolytically derived peptides by measuring the levels of alpha-soluble amyloid precursor protein, total soluble amyloid precursor proteins (alpha- and beta-soluble amyloid precursor protein combined) and beta-amyloid in cerebrospinal fluid from three-, 13- and 23-month-old Fischer-344 rats. Western blot analysis using selective antibodies revealed 50% less total soluble amyloid precursor protein and a-soluble amyloid precursor protein in cisternal cerebrospinal fluid from 23-month-old rats compared with three- and 13-month-old animals. Mass spectrometric analysis indicated, however, that beta-amyloid in cerebrospinal fluid was not different between the three age groups. In a second group of young (five to six months of age) and aged (24-25 months of age) rats, spatial working and reference memory were assessed in a water maze followed by collection of cerebrospinal fluid. As a group, the aged rats consistently performed below the young rats in both working and reference memory tests. The aged rats also had 49% less cerebrospinal fluid alpha-soluble amyloid precursor protein than did their younger counterparts. There was a positive correlation (r= 0.52-0.57, P < 0.001) between performance in spatial memory tasks and cerebrospinal fluid alpha-soluble amyloid precursor protein in these young and aged rats. These results suggest that there is a positive association between cerebrospinal fluid levels of alpha-soluble amyloid precursor protein and cognitive performance in rats, and that alpha-soluble amyloid precursor protein may be involved in the spatial learning and memory changes that accompany ageing.
Assuntos
Envelhecimento/metabolismo , Peptídeos beta-Amiloides/líquido cefalorraquidiano , Precursor de Proteína beta-Amiloide/líquido cefalorraquidiano , Endopeptidases/metabolismo , Transtornos da Memória/metabolismo , Fragmentos de Peptídeos/líquido cefalorraquidiano , Secretases da Proteína Precursora do Amiloide , Peptídeos beta-Amiloides/análise , Precursor de Proteína beta-Amiloide/análise , Animais , Córtex Cerebral/química , Córtex Cerebral/metabolismo , Cognição/fisiologia , Condicionamento Psicológico , Hipocampo/química , Hipocampo/metabolismo , Masculino , Aprendizagem em Labirinto , Fragmentos de Peptídeos/análise , Ratos , Ratos Endogâmicos F344 , Retenção Psicológica , Solubilidade , Percepção Espacial/fisiologia , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Acuidade VisualAssuntos
Doença de Alzheimer/genética , Precursor de Proteína beta-Amiloide/genética , Cromossomos Artificiais de Levedura , Proteínas de Membrana/genética , Precursor de Proteína beta-Amiloide/metabolismo , Animais , Humanos , Camundongos , Camundongos Transgênicos , Mutação , Presenilina-1 , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
The insulin-like growth factor binding proteins (IGFBPs) act to modulate the growth and differentiation of the gastrointestinal mucosa by regulating cellular responses to the important mitogens, the insulin-like growth factors (IGFs). The transplanted small bowel must maintain the normal growth factor interrelationships and signaling pathways despite the potential of host rejection. Ostomy effluent of patients after small bowel or combined liver/small bowel transplantation was assayed for IGFBPs to investigate the effect of rejection on the IGF-IGFBP axis. Seventeen patients were studied over an 18-month period. The transplanted small bowel produced no measurable IGFBPs in the ostomy effluent under normal circumstances. However, when rejection was taking place the ostomy effluent was found to have measurable IGFBP levels in 6 of 12 episodes, the 6 episodes occurring in 6 different patients. Mostly, the IGFBPs present did not exhibit a serum-like pattern indicating the secretion of IGFBPs into the effluent was not the result of loss of mucosal barrier integrity. There were no statistically significant differences in protein content of the ostomy fluids in the presence or absence of rejection. Our results suggest that the gastrointestinal IGF axis is altered during some instances of transplanted small bowel rejection, with increased secretion of IGFBPs.
Assuntos
Proteínas de Ligação a Fator de Crescimento Semelhante a Insulina/metabolismo , Mucosa Intestinal/metabolismo , Intestino Delgado/transplante , Adolescente , Adulto , Líquidos Corporais/metabolismo , Criança , Pré-Escolar , Feminino , Humanos , Lactente , Transplante de Fígado/fisiologia , MasculinoRESUMO
From March 1994 to February 1996, 28 infertile couples participated in the oocyte donation program in 33 treatment cycles at the Southeastern Fertility Institute. Of the 31 cycles with embryo transfer, 15 cycles (48.4 percent) resulted in a clinical pregnancy with fetal heart beat by ultrasound. The spontaneous first trimester abortion rate was 3/15 (20 percent), multiple pregnancy rate 3/15 (20 percent), live birth rate 11/15 (73.3 percent) and delivery rate 12/15 (80 percent). It is recommended that oocyte donation procedure is a highly successful treatment option for women with ovarian failure or repeated unsuccessful trials of assisted reproductive technologies.
Assuntos
Infertilidade Feminina , Doação de Oócitos , Adulto , Transferência Embrionária , Feminino , Humanos , South Carolina , Resultado do TratamentoRESUMO
Missense mutations in the beta-amyloid precursor protein gene (APP) co-segregate with a small subset of autosomal dominant familial Alzheimer's disease (FAD) cases wherein deposition of the 39-43 amino acid beta-amyloid (A beta) peptide and neurodegeneration are principal neuropathological hallmarks. To accurately examine the effect of missense mutations on APP metabolism and A beta production in vivo, we have introduced yeast artificial chromosomes (YACs) containing the entire approximately 400 kbp human APP gene encoding APP harboring either the asparagine for lysine and leucine for methionine FAD substitution at codons 670 and 671 (APP(K670N/M671L)), the isoleucine for valine FAD substitution at codon 717 (APP(V7171)) or a combination of both substitutions into transgenic mice. We demonstrate that, relative to YAC transgenic mice expressing wild-type APP, high levels of A beta peptides are detected in the brains of YAC transgenic mice expressing human APP(K670N/M671L) that is associated with a concomitant diminution in the levels of apha-secretase-generated soluble APP derivatives. Moreover, the levels of longer A beta peptides (species terminating at amino acids 42/43) are elevated in YAC transgenic mice expressing human APP(V7171). These mice should prove valuable for detailed analysis of the in vivo effects of the APP FAD mutations in a variety of tissues and throughout aging and for testing therapeutic agents that specifically alter APP metabolism and A beta production.
Assuntos
Doença de Alzheimer/genética , Precursor de Proteína beta-Amiloide/metabolismo , Cromossomos Artificiais de Levedura , Precursor de Proteína beta-Amiloide/genética , Animais , Técnicas de Cultura de Células , Expressão Gênica , Vetores Genéticos , Humanos , Camundongos , Camundongos Transgênicos/genética , Mutagênese , Reação em Cadeia da Polimerase , RNA Mensageiro/metabolismo , Transgenes/genéticaRESUMO
OBJECTIVE: Our purpose was to evaluate a tactile non-ultrasonographically directed approach to transvaginal tubal cannulation and embryo transfer during the zygote intrafallopian tube transfer procedure. STUDY DESIGN: We studied the application of the tactile approach to transvaginal tubal cannulation and embryo transfer in 68 consecutive zygote intrafallopian tube transfer cycles. RESULTS: The tactile approach to transvaginal tubal cannulation and subsequent embryo transfer was successful in 93% of couples. Pregnancy occurred in 16 of the 63 (25%) women undergoing a successful transvaginal embryo transfer. CONCLUSION: The tactile approach to transvaginal tubal cannulation was found to be easily learned and provided a safe and convenient method of tubal embryo transfer. This low-cost approach, which has application to transfer of both embryos and gametes, deserves further study.
Assuntos
Cateterismo/métodos , Transferência Intratubária do Zigoto/métodos , Cateterismo/instrumentação , Estudos de Avaliação como Assunto , Feminino , Humanos , Gravidez , Tato , Resultado do Tratamento , Vagina , Transferência Intratubária do Zigoto/estatística & dados numéricosAssuntos
Doenças Peritoneais/prevenção & controle , Complicações Pós-Operatórias/prevenção & controle , Aderências Teciduais/prevenção & controle , Anti-Inflamatórios/uso terapêutico , Anticoagulantes/uso terapêutico , Infecções Bacterianas/prevenção & controle , Dextranos/uso terapêutico , Feminino , Doenças dos Genitais Femininos/cirurgia , Humanos , Próteses e Implantes , Terapia TrombolíticaRESUMO
The use of a digital audio-video mixer is advocated when concurrent endoscopic procedures, such as laparoscopy and hysteroscopy, are performed. The device can also assist in postproduction work.
Assuntos
Gravação de Videoteipe/instrumentação , Desenho de Equipamento , Estudos de Avaliação como Assunto , Humanos , Histeroscópios , LaparoscópiosRESUMO
We have described a 28-year-old diabetic woman who had necrotizing fasciitis of the perineum three years after receiving a living related renal transplant. The diagnosis of necrotizing fasciitis was made early and she was referred to a tertiary care center where she received radical perineal debridement and aggressive medical and surgical follow-up. Necrotizing fasciitis in a transplant patient is rare; review of the literature shows few cases and no survivors. Our patient has returned to a normal life despite continuation of all immunosuppressive therapy throughout the entire hospital course. In addition, she had a good cosmetic result despite the large necrotic perineal infection. Her survival can be attributed to early diagnosis and referral, immediate and extensive debridement, and aggressive protein replacement.
Assuntos
Fasciite/diagnóstico , Transplante de Rim , Períneo , Complicações Pós-Operatórias/diagnóstico , Adulto , Desbridamento , Fasciite/patologia , Fasciite/cirurgia , Fasciite/terapia , Feminino , Hidratação , Humanos , Necrose , Complicações Pós-Operatórias/patologia , Complicações Pós-Operatórias/cirurgia , Complicações Pós-Operatórias/terapia , ReoperaçãoRESUMO
Hamster tracheal explants were maintained in culture in the presence or absence of benzo[a]pyrene (BP), crocidolite asbestos, or BP-coated crocidolite. Dose-dependent squamous metaplasia was observed in the treated samples. L-Ascorbic acid and DL-alpha-tocopherol were able to partially protect the tracheal explants from the metaplastic response induced by crocidolite. Furthermore, ascorbic acid reduced the extent of metaplasia observed in hamster tracheal explants that were exposed to BP-crocidolite.
Assuntos
Amianto/toxicidade , Ácido Ascórbico/farmacologia , Benzo(a)pireno/toxicidade , Traqueia/efeitos dos fármacos , Animais , Cricetinae , Masculino , Mesocricetus , Metaplasia/prevenção & controle , Traqueia/patologiaRESUMO
Two hundred eighty-five charts were reviewed from patients who underwent surgery for ectopic pregnancy. Excluded were patients with previous tubal reparative surgery, linear salpingotomy, or failed sterilization. The incidence of isthmic ectopic pregnancy in the remaining 255 cases was 15.3%. The association of salpingitis isthmica nodosa (SIN) and isthmic ectopic pregnancy was determined by review of resected tubal segments. SIN was noted in 17 of 37 cases (45.9%) of isthmic ectopic pregnancy. SIN places the patient at risk for recurrent ectopic pregnancy or infertility. Recommended conservative management of isthmic ectopic pregnancy is segmental resection with postoperative emphasis on documentation of SIN when present. Postoperative hysterosalpingography is recommended with an abnormal contralateral tube or when SIN is noted in the resected tubal segment. Management options after an isthmic ectopic pregnancy when future fertility is desired are presented.
Assuntos
Gravidez Tubária/etiologia , Salpingite/complicações , Tubas Uterinas/cirurgia , Feminino , Humanos , Gravidez , Gravidez Tubária/cirurgiaRESUMO
The fertilization rates of mature oocytes during in vitro fertilization and embryo transfer (IVF-ET) using fetal cord serum-supplemented insemination media were greater than or equal to 57% for five infertile couples without sperm antibodies (group 1). But they were less than or equal to 50% for four of nine infertile couples (group 2) with cytotoxic sperm antibodies in both partners (n = 6) or the husband alone (n = 3). Two women in group 1 were successful in achieving normal, full-term pregnancies with the delivery of normal infants (chi2 = 4.2, P less than 0.05, by chi-square analysis). One of them consistently tested negative for sperm antibodies, while her husband was previously treated with antibiotics for infection and transient sperm antibodies in the seminal plasma. Subsequently, antibody titers in the husband were in the normal range when the successful IVF-ET was performed. One woman in group 2, with antibodies to her autoimmune husband's sperm but not control sperm and with a long-standing poor postcoital test sperm motility, conceived through artificial insemination with donor sperm (AID) after failing to conceive with her husband through IVF-ET. These data suggest that the presence of cytotoxic sperm antibodies in the serum and/or secretions of both partners reduces the rates of fertilization of mature oocytes in spite of using fetal cord serum in the IVF media. Pregnancy achievement is impaired in this group.