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Fundamental to holobiont biology is recognising how variation in microbial composition and function relates to host phenotypic variation. Sponges often exhibit considerable phenotypic plasticity and also harbour dense microbial communities that function to protect and nourish hosts. One of the most prominent sponge genera on Caribbean coral reefs is Agelas. Using a comprehensive set of morphological (growth form, spicule), chemical and molecular data on 13 recognised species of Agelas in the Caribbean basin, we were able to define only five species (=clades) and found that many morphospecies designations were incongruent with phylogenomic and population genetic analyses. Microbial communities were also strongly differentiated between phylogenetic species, showing little evidence of cryptic divergence and relatively low correlation with morphospecies assignment. Metagenomic analyses also showed strong correspondence to phylogenetic species, and to a lesser extent, geographical and morphological characters. Surprisingly, the variation in secondary metabolites produced by sponge holobionts was explained by geography and morphospecies assignment, in addition to phylogenetic species, and covaried significantly with a subset of microbial symbionts. Spicule characteristics were highly plastic, under greater impact from geographical location than phylogeny. Our results suggest that while phenotypic plasticity is rampant in Agelas, morphological differences within phylogenetic species affect functionally important ecological traits, including the composition of the symbiotic microbial communities and metabolomic profiles.
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Agelas , Poríferos , Animales , Filogenia , Región del Caribe , Indias Occidentales , Arrecifes de Coral , Poríferos/genéticaRESUMEN
BACKGROUND AND PURPOSE: Cerebrospinal fluid (CSF) kappa free light chains (FLCs) may be a more sensitive marker of intrathecal immunoglobulin (Ig)G synthesis compared with oligoclonal bands (OCBs). Our aim was to retrospectively determine the additional value of the kappa and lambda index (CSF FLC/serum FLC)/(CSF albumin/serum albumin) in predicting a multiple sclerosis (MS) diagnosis in a group of OCB-negative patients with suspected MS. METHODS: The CSF and serum kappa and lambda FLCs were tested using the Freelite kit (serum) and Freelite Mx (CSF) assay (The Binding Site Group, Bimingham, UK) in 391 OCB-negative patients with suspected/possible MS and in 54 OCB-positive patients with MS. RESULTS: The CSF kappa FLC levels were below the detection limit (0.27 mg/L) in 61% of patients. Using quantitative data, we found the best kappa index cut-off value for the prediction of MS to be 5.8. A kappa index ≥5.8 was present in 25% of OCB-negative MS (23/92) and in 98% of OCB-positive patients with MS. Using a qualitative approach and a kappa index cut-off of 5.9, based on literature data, we likewise found that 24% of OCB-negative patients with MS had a kappa index ≥5.9, compared with 5.4% of OCB-negative patients without MS (P < 0.001). No reliable data could be obtained for the lambda index; lambda FLCs were below the detection limit (0.68 mg/L) in 90% of CSF samples. CONCLUSIONS: The kappa index could contribute to the identification of OCB-negative patients with a high probability of an MS diagnosis. Using more sensitive techniques might even improve the diagnostic performance of the kappa index and better define the role of the lambda index.
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Inmunoglobulina G/líquido cefalorraquídeo , Cadenas kappa de Inmunoglobulina/líquido cefalorraquídeo , Cadenas lambda de Inmunoglobulina/líquido cefalorraquídeo , Esclerosis Múltiple/diagnóstico , Bandas Oligoclonales/líquido cefalorraquídeo , Adulto , Biomarcadores/líquido cefalorraquídeo , Femenino , Humanos , Masculino , Persona de Mediana Edad , Esclerosis Múltiple/líquido cefalorraquídeo , Estudios RetrospectivosRESUMEN
BACKGROUND AND PURPOSE: Autoimmune encephalitides (AE) include a spectrum of neurological disorders whose diagnosis revolves around the detection of neuronal antibodies (Abs). Consensus-based diagnostic criteria (AE-DC) allow clinic-serological subgrouping of AE, with unclear prognostic implications. The impact of AE-DC on patients' management was studied, focusing on the subgroup of Ab-negative-AE. METHODS: This was a retrospective multicenter study on patients fulfilling AE-DC. All patients underwent Ab testing with commercial cell-based assays (CBAs) and, when available, in-house assays (immunohistochemistry, live/fixed CBAs, neuronal cultures) that contributed to defining final categories. Patients were classified as Ab-positive-AE [N-methyl-d-aspartate-receptor encephalitis (NMDAR-E), Ab-positive limbic encephalitis (LE), definite-AE] or Ab-negative-AE (Ab-negative-LE, probable-AE, possible-AE). RESULTS: Commercial CBAs detected neuronal Abs in 70/118 (59.3%) patients. Testing 37/48 Ab-negative cases, in-house assays identified Abs in 11 patients (29.7%). A hundred and eighteen patients fulfilled the AE-DC, 81 (68.6%) with Ab-positive-AE (Ab-positive-LE, 40; NMDAR-E, 32; definite-AE, nine) and 37 (31.4%) with Ab-negative-AE (Ab-negative-LE, 17; probable/possible-AE, 20). Clinical phenotypes were similar in Ab-positive-LE versus Ab-negative-LE. Twenty-four/118 (20.3%) patients had tumors, and 19/118 (16.1%) relapsed, regardless of being Ab-positive or Ab-negative. Ab-positive-AE patients were treated earlier than Ab-negative-AE patients (P = 0.045), responded more frequently to treatments (92.3% vs. 65.6%, P < 0.001) and received second-line therapies more often (33.3% vs. 10.8%, P = 0.01). Delays in first-line therapy initiation were associated with poor response (P = 0.022; odds ratio 1.02; confidence interval 1.00-1.04). CONCLUSIONS: In-house diagnostics improved Ab detection allowing better patient management but was available in a patient subgroup only, implying possible Ab-positive-AE underestimation. Notwithstanding this limitation, our findings suggest that Ab-negative-AE and Ab-positive-AE patients share similar oncological profiles, warranting appropriate tumor screening. Ab-negative-AE patients risk worse responses due to delayed and less aggressive treatments.
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Encefalitis/diagnóstico , Enfermedad de Hashimoto/diagnóstico , Neuronas/inmunología , Fenotipo , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Niño , Preescolar , Encefalitis/inmunología , Femenino , Enfermedad de Hashimoto/inmunología , Humanos , Inmunohistoquímica , Lactante , Masculino , Persona de Mediana Edad , Receptores de N-Metil-D-Aspartato/inmunología , Estudios Retrospectivos , Adulto JovenRESUMEN
BACKGROUND AND PURPOSE: Acquired neuromyotonia can occur in patients with thymoma, alone or in association with myasthenia gravis (MG), but the clinical prognostic significance of such comorbidity is largely unknown. The clinico-pathological features were investigated along with the occurrence of neuromyotonia as predictors of tumour recurrence in patients with thymoma-associated myasthenia. METHODS: A total number of 268 patients with thymomatous MG were studied retrospectively. Patients with symptoms of spontaneous muscle overactivity were selected for autoantibody testing using immunohistology for neuronal cell-surface proteins and cell-based assays for contactin-associated protein 2 (CASPR2), leucine-rich glioma inactivated 1 (LGI1), glycine receptor and Netrin-1 receptor antibodies. Neuromyotonia was diagnosed according to the presence of typical electromyography abnormalities and/or autoantibodies against LGI1/CASPR2. RESULTS: Overall, 33/268 (12%) MG patients had a thymoma recurrence. Five/268 (2%) had neuromyotonia, four with typical autoantibodies, including LGI1 (n = 1), CASPR2 (n = 1) or both (n = 2). Three patients had Netrin-1 receptor antibodies, two with neuromyotonia and concomitant CASPR2+LGI1 antibodies and one with spontaneous muscle overactivity without electromyography evidence of neuromyotonia. Thymoma recurrence was more frequent in those with (4/5, 80%) than in those without (28/263, 10%, P < 0.001) neuromyotonia. Neuromyotonia preceded the recurrence in 4/5 patients. In univariate analysis, predictors of thymoma recurrence were age at thymectomy [odds ratio (OR) 0.95, 95% confidence interval (CI) 0.93-0.97], Masaoka stage ≥IIb (OR 10.73, 95% CI 2.38-48.36) and neuromyotonia (OR 41.78, 95% CI 4.71-370.58). CONCLUSIONS: De novo occurrence of neuromyotonia in MG patients with previous thymomas is a rare event and may herald tumour recurrence. Neuronal autoantibodies can be helpful to assess the diagnosis. These observations provide pragmatic risk stratification for tumour vigilance in patients with thymomatous MG.
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Síndrome de Isaacs/complicaciones , Miastenia Gravis/complicaciones , Timoma/complicaciones , Neoplasias del Timo/complicaciones , Adulto , Autoanticuerpos/sangre , Electromiografía , Femenino , Humanos , Masculino , Proteínas de la Membrana/inmunología , Persona de Mediana Edad , Miastenia Gravis/sangre , Recurrencia Local de Neoplasia , Netrina-1/inmunología , Estudios Retrospectivos , Timoma/sangre , Neoplasias del Timo/sangreRESUMEN
We describe two patients with mitochondrial DNA mutations in the gene encoding cytochrome b (m.15579A>G, p.Tyr278Cys and m.15045G>A p.Arg100Gln), which presented as a pure myopathic form (exercise intolerance), with an onset in childhood. Diagnosis was delayed, because acylcarnitine profile showed an increase in medium and long-chain acylcarnitines, suggestive of multiple acyl-CoA dehydrogenase deficiency, riboflavin transporter deficiency or FAD metabolism disorder. Implication of cytochrome b in fatty acid oxidation, and physiopathology of the mutations are discussed.
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Citocromos b/genética , Deficiencia Múltiple de Acil Coenzima A Deshidrogenasa/diagnóstico , Enfermedades Musculares/diagnóstico , Enfermedades Musculares/genética , Mutación Missense , Adulto , Anciano , ADN Mitocondrial/genética , Diagnóstico Diferencial , Tolerancia al Ejercicio/genética , Humanos , Masculino , Deficiencia Múltiple de Acil Coenzima A Deshidrogenasa/genéticaRESUMEN
Introduction: Acute amnestic syndrome is an uncommon clinical presentation of neurological disease. Differential diagnosis encompasses several syndromes including Wernicke-Korsakoff and transient global amnesia (TGA). Structural lesions of the fornix account for a minority of cases of acute amnestic syndromes. Etiology varies from iatrogenic injury to ischemic, inflammatory, or neoplastic lesions. A prompt diagnosis of the underlying pathology is essential but challenging. The aim of this review is to systematically review the existing literature regarding cases of acute amnestic syndrome associated with non-iatrogenic lesions of the fornix. Methods: We performed a systematic literature search on PubMed, Scopus, and Web of Science up to September 2023 to identify case reports and case series of patients with amnestic syndrome due to fornix lesions. The systematic review was conducted according to PRISMA guidelines. The research was limited to articles written in English. Cases of fornix damage directly ascribable to a surgical procedure were excluded. Results: A total of 52 publications reporting 55 cases were included in the review. Focusing on acute/subacute onset, vascular etiology was highly prevalent, being responsible for 78% of cases, 40/55 (74%) of which were due to acute ischemic stroke. The amnestic syndrome was characterized by anterograde amnesia in all patients, associated with retrograde amnesia in 27% of cases. Amnesia was an isolated presentation in most cases. Up to two thirds of patients had persistent memory deficits of any severity at follow-up. Discussion: Acute amnestic syndrome can be rarely caused by fornix lesions. In most cases of acute/subacute presentation, the etiology is ischemic stroke, mainly caused by strokes involving the subcallosal artery territory. The differential diagnosis is challenging and a distinction from common mimics is often difficult on a clinical basis. A high index of suspicion should be maintained to avoid misdiagnosis and provide adequate acute treatment to patients with time-dependent disease, also employing advanced neuroimaging. More research is needed to better understand the outcome and identify prognostic factors in patients with amnestic syndrome due to fornix lesions.
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BACKGROUND: Myelin oligodendrocyte glycoprotein antibody-associated encephalomyelitis (MOG-EM; also termed MOG antibody-associated disease, MOGAD) is the most important differential diagnosis of both multiple sclerosis and neuromyelitis optica spectrum disorders. A recent proposal for new diagnostic criteria for MOG-EM/MOGAD explicitly recommends the use of immunoglobulin G subclass 1 (IgG1)- or IgG crystallizable fragment (Fc) region-specific assays and allows the use of heavy-and-light-chain-(H+L) specific assays for detecting MOG-IgG. By contrast, the utility of MOG-IgG3-specific testing has not been systematically evaluated. OBJECTIVE: To assess whether the use of MOG-IgG3-specific testing can improve the sensitivity of MOG-IgG testing. METHODS: Re-testing of 22 patients with a definite diagnosis of MOG-EM/MOGAD and clearly positive MOG-IgG status initially but negative or equivocal results in H+L- or Fc-specific routine assays later in the disease course (i.e. patients with spontaneous or treatment-driven seroreversion). RESULTS: In accordance with previous studies that had used MOG-IgG1-specific assays, IgG subclass-specific testing yielded a higher sensitivity than testing by non-subclass-specific assays. Using subclass-specific secondary antibodies, 26/27 supposedly seroreverted samples were still clearly positive for MOG-IgG, with MOG-IgG1 being the most frequently detected subclass (25/27 [93%] samples). However, also MOG-IgG3 was detected in 14/27 (52%) samples (from 12/22 [55%] patients). Most strikingly, MOG-IgG3 was the predominant subclass in 8/27 (30%) samples (from 7/22 [32%] patients), with no unequivocal MOG-IgG1 signal in 2 and only a very weak concomitant MOG-IgG1 signal in the other six samples. By contrast, no significant MOG-IgG3 reactivity was seen in 60 control samples (from 42 healthy individuals and 18 patients with MS). Of note, MOG-IgG3 was also detected in the only patient in our cohort previously diagnosed with MOG-IgA+/IgG- MOG-EM/MOGAD, a recently described new disease subvariant. MOG-IgA and MOG-IgM were negative in all other patients tested. CONCLUSIONS: In some patients with MOG-EM/MOGAD, MOG-IgG is either exclusively or predominantly MOG-IgG3. Thus, the use of IgG1-specific assays might only partly overcome the current limitations of MOG-IgG testing and-just like H+L- and Fcγ-specific testing-might overlook some genuinely seropositive patients. This would have potentially significant consequences for the management of patients with MOG-EM/MOGAD. Given that IgG3 chiefly detects proteins and is a strong activator of complement and other effector mechanisms, MOG-IgG3 may be involved in the immunopathogenesis of MOG-EM/MOGAD. Studies on the frequency and dynamics as well as the clinical and therapeutic significance of MOG-IgG3 seropositivity are warranted.
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Autoanticuerpos , Inmunoglobulina G , Glicoproteína Mielina-Oligodendrócito , Humanos , Glicoproteína Mielina-Oligodendrócito/inmunología , Inmunoglobulina G/sangre , Inmunoglobulina G/inmunología , Femenino , Masculino , Adulto , Persona de Mediana Edad , Autoanticuerpos/sangre , Sensibilidad y Especificidad , Adulto Joven , Anciano , Encefalomielitis/diagnóstico , Encefalomielitis/inmunología , Encefalomielitis/sangreRESUMEN
BACKGROUND: Myelin oligodendrocyte glycoprotein antibody-related spectrum disorders (MOG-SD) are a heterogeneous group of inflammatory demyelinating diseases of the central nervous system, usually responsive to conventional immunosuppressive therapies. However, knowledge about treatment of non-responder patients is scarce. METHODS: We report on a 20-year-old MOG-SD patient who experienced clinical deterioration despite rituximab-induced B-cell depletion. RESULTS: Rescue therapy with tocilizumab (TCZ) prevented further relapses, with reduction of spinal-cord load on MRI, and a remarkable reduction of disability at the two-year follow-up. CONCLUSION: Our observations suggest that TCZ could induce clinico-radiologic improvements, which make it as an option for the treatment of MOG-SD.
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Anticuerpos Monoclonales Humanizados/uso terapéutico , Enfermedades Autoinmunes Desmielinizantes SNC/tratamiento farmacológico , Factores Inmunológicos/uso terapéutico , Glicoproteína Mielina-Oligodendrócito/inmunología , Enfermedades Autoinmunes Desmielinizantes SNC/complicaciones , Enfermedades Autoinmunes Desmielinizantes SNC/diagnóstico por imagen , Enfermedades Autoinmunes Desmielinizantes SNC/patología , Humanos , Masculino , Neuritis Óptica/complicaciones , Rituximab/uso terapéutico , Médula Espinal/diagnóstico por imagen , Médula Espinal/efectos de los fármacos , Resultado del Tratamiento , Adulto JovenRESUMEN
Rituximab is efficacious in myelin-associated glycoprotein (MAG) polyneuropathy, but the question on timing of retreatments is open. We studied 21 anti-MAG polyneuropathy patients who responded to a first cycle of rituximab, were followed-up for an average of 11.2â¯years, and were retreated only when relapsing. Baseline serum B-cell-activating factor (BAFF) levels were measured. Clinical improvements lasted on average 6â¯years, and as many as 71% of the patients resulted long-lasting responders. Severity of disease and high serum BAFF levels (cut-off ≥860â¯pg/mL for relapse risk) at onset seemed to predict worse prognosis. Measurements of these variables could help deal with the issue of maintenance rituximab therapy in MAG polyneuropathy.
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Autoanticuerpos/sangre , Factores Inmunológicos/administración & dosificación , Glicoproteína Asociada a Mielina/sangre , Polineuropatías/sangre , Polineuropatías/tratamiento farmacológico , Rituximab/administración & dosificación , Adulto , Anciano , Anciano de 80 o más Años , Autoanticuerpos/inmunología , Femenino , Estudios de Seguimiento , Humanos , Masculino , Persona de Mediana Edad , Glicoproteína Asociada a Mielina/inmunología , Polineuropatías/inmunología , Factores de Tiempo , Resultado del TratamientoRESUMEN
Although estrogen receptor (ER)-alpha is expressed in both benign and malignant ovarian tumors, the role of ER in ovarian carcinogenesis of epithelial tumors is still unknown. In view of the recent characterization of ER-beta, a second form of ER that seems to be highly expressed in ovaries, we reexamined this issue by studying the relative expression of ER-alpha and -beta in human ovarian tumor progression. We developed a competitive PCR assay based on coamplification of the two ERs in target nucleotide sequences displaying a high homology (exons 3 and 4). Coamplification experiments with varying amounts of plasmids containing ER-alpha and -beta cDNAs showed that this assay was reliable for discriminating as little as a 2-fold difference in the initial ER-alpha:ER-beta cDNA ratio. The relative expression of ER-alpha compared with ER-beta mRNAs was studied in human ovarian cancer cell lines (n = 5) and in normal ovaries (n = 6), then in human benign and malignant tumor samples including ovarian cysts (n = 24), borderline tumors (n = 3), and cancers (n = 10). In normal ovaries, ER-beta mRNA was the predominant ER form, whereas in ovarian cancer cell lines ER-alpha mRNA was markedly increased as compared with ER-beta. In benign and borderline tumors, ER-beta mRNA was detected in 78% of tumors, whereas ER-alpha mRNA was detected in 29%. In ovarian carcinomas, both ER-alpha and -beta mRNAs were expressed in 80% of tumors. The ER-alpha:ER-beta mRNA ratio was >1 in only one cyst sample (4%). In contrast, the ER-alpha:ER-beta mRNA ratio was markedly increased in ovarian cancers because 60% showed an ER-alpha:ER-beta mRNA >1. In situ hybridization experiments showed overlapping tissular distribution of ER-beta and -alpha expression in cancers and cysts, with a main localization in the epithelium and only a low level of expression in stromal cells. In summary, we found an increase in the ER-alpha:ER-beta mRNA ratio in ovarian carcinomas as compared with normal ovaries and cysts. These data suggest that overexpression of ER-alpha relative to ER-beta mRNA may be a marker of ovarian carcinogenesis.
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Biomarcadores de Tumor/biosíntesis , Neoplasias Ováricas/metabolismo , ARN Mensajero/biosíntesis , Receptores de Estrógenos/biosíntesis , ADN Complementario/genética , ADN Complementario/metabolismo , Progresión de la Enfermedad , Receptor alfa de Estrógeno , Receptor beta de Estrógeno , Femenino , Humanos , Hibridación in Situ , Enfermedades del Ovario/metabolismo , Enfermedades del Ovario/patología , Neoplasias Ováricas/patología , Neoplasias Ováricas/ultraestructura , Plásmidos , Isoformas de Proteínas , Receptores de Estrógenos/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Células Tumorales CultivadasRESUMEN
Combined central and peripheral demyelination (CCPD) is rare, and current knowledge is based on case reports and small case series. The aim of our study was to describe the clinical features, diagnostic results, treatment and outcomes in a large cohort of patients with CCPD. Thirty-one patients entered this retrospective, observational, two-center study. In 20 patients (65%) CCPD presented, after an infection, as myeloradiculoneuropathy, encephalopathy, cranial neuropathy, length-dependent peripheral neuropathy, or pseudo-Guillain-Barré syndrome. Demyelinating features of peripheral nerve damage fulfilling European Federation of Neurological Societies/Peripheral Nerve Society (EFNS/PNS) electrodiagnostic criteria for CIDP were found in 23 patients (74%), and spatial dissemination of demyelinating lesions on brain MRI fulfilling the 2010 McDonald criteria for multiple sclerosis (MS) in 11 (46%). Two thirds of the patients had a relapsing or progressive disease course, usually related to the appearance of new spinal cord lesions or worsening of the peripheral neuropathy, and showed unsatisfactory responses to high-dose corticosteroids and intravenous immunoglobulins. The clinical presentation of CCPD was severe in 22 patients (71%), who were left significantly disabled. Our data suggest that CCPD has heterogeneous features and shows frequent post-infectious onset, primary peripheral nervous system or central nervous system involvement, a monophasic or chronic disease course, inadequate response to treatments, and a generally poor outcome. We therefore conclude that the current diagnostic criteria for MS and CIDP may not fully encompass the spectrum of possible manifestations of CCPD, whose pathogenesis remains largely unknown.
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Enfermedades Desmielinizantes/diagnóstico por imagen , Enfermedades Desmielinizantes/terapia , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Estudios de Cohortes , Femenino , Estudios de Seguimiento , Humanos , Imagen por Resonancia Magnética , Masculino , Persona de Mediana Edad , Polirradiculoneuropatía/diagnóstico por imagen , Polirradiculoneuropatía/terapia , Polirradiculoneuropatía Crónica Inflamatoria Desmielinizante/diagnóstico por imagen , Polirradiculoneuropatía Crónica Inflamatoria Desmielinizante/terapia , Estudios Retrospectivos , Resultado del Tratamiento , Adulto JovenRESUMEN
Most recently reported methods to select early hematopoietic cells basically rely on the depletion of committed progenitors. This task is generally accomplished by laborious procedures, which are sometimes difficult to reproduce. To simplify the selection method, we took advantage of the expression of the transferrin receptor (CD71) by proliferating committed progenitors and the lack of CD71 on noncycling immature progenitors. A monoclonal antibody (MAB) reactive with CD71 has been conjugated to the Saponaria officinalis seed ribosome-inactivating protein (SO6). The immunotoxin (IT) complex was used at increasing concentrations on normal non-phagocytizing bone marrow cells. A complete and reproducible killing effect on myeloid (colony-forming unit-granulocyte/macrophage [CFU-GM]) and erythroid (burst-forming unit-erythroid [BFU-E]) progenitors was observed for IT concentrations of 1 x 10(-7) M. Unconjugated SO6 or anti-CD71 MAB had no effect on cell growth and viability. IT-resistant cells were able to generate CFU-GM after 7, 14, and 21 days of suspension culture in the presence of 5637 CM. Maximal CFU-GM values were obtained at day 21 and nearly approached the pretreatment values (mean 2587 vs. 3877 CFU-GM/mL). Growth factor enhancement of CFU-GM yield was obtained only by stem cell factor (SCF) at day 7; SCF, as well as granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin-3 (IL-3), had an enhancing effect at days 14 and 21. IT toxicity on highly immature progenitors was ruled out by evaluating the growth of long-term culture-initiating cells (LTC-IC) from IT-treated cultures. LTC-IC frequency was found to be 1 out of 1506 seeded cells, which is within the range of normal untreated BM cells. In conclusion, anti-CD71 IT allows a simple and complete depletion of committed progenitors while sparing immature hematopoietic cells. The high CD71 expression by leukemic cells makes the procedure potentially suitable for in vitro purging.
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Antígenos CD/inmunología , Antígenos de Diferenciación de Linfocitos B/inmunología , Separación Celular , Células Madre Hematopoyéticas/citología , Inmunotoxinas/farmacología , N-Glicosil Hidrolasas , Proteínas de Plantas/farmacología , Anticuerpos Monoclonales , Muerte Celular , División Celular , Células Cultivadas , Células Precursoras Eritroides/citología , Factor Estimulante de Colonias de Granulocitos y Macrófagos/farmacología , Granulocitos/citología , Factores de Crecimiento de Célula Hematopoyética/farmacología , Interleucina-3/farmacología , Cinética , Macrófagos/citología , Proteínas de Plantas/administración & dosificación , Receptores de Transferrina , Proteínas Inactivadoras de Ribosomas Tipo 1 , Saporinas , Factor de Células MadreRESUMEN
Bile salt-dependent lipase (BSDL), an enzyme normally found in human pancreatic secretions is a 100 kDa glycoprotein. A BSDL-specific 477 bp cDNA probe was prepared by performing polymerase chain reaction experiments. This cDNA was used to probe mRNAs extracted from human pancreatic tissue and tumoral cell lines. Two mRNAs were detected in normal human pancreas at 2.2 and 1.3 to 1.5 kb. In human pancreatic tumoral cells, mRNAs encoding for the BSDL were detected using in situ hybridization, and proteins with an M(r) of 46,000 to 48,000 were translated into an in vitro system using mRNAs extracted from these cells. Using an immunoprecipitation procedure, we observed here that the specific BSDL polyclonal antibodies recognized three proteins of 100 +/- 5 (p100), 46 +/- 2 (p46) and 22.7 +/- 1.2 (p23) kDa, respectively in the soluble extracts of normal adult human pancreas. The p100 protein was probably the glycosylated product resulting from the translation of the 2.2 kb transcript. The p46 protein, which electrophoresed as a doublet was the main component immunoprecipitated from extract of a differentiated human pancreatic adenocarcinoma as well as from the extracts of two pancreatic cell lines, BxPC-3 and SOJ-6. In addition, the p46 immunoform of the BSDL was detected in cell-free medium from SOJ-6 cell line and its expression was found to be correlated with the secretion of an esterolytic activity on 4-nitrophenyl caproate, whereas the BxPC-3 cell line neither secreted the p46 nor showed any esterolytic activity on this substrate. The p46 may be either a short variant of BSDL resulting from the translation of the 1.3 to 1.5 kb transcript or a protein structurally related to the enzyme. The p46 doublet immunoform was detected in the human pancreatic secretion.
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Lipasa/análisis , Proteínas de Neoplasias/análisis , Neoplasias Pancreáticas/química , Esterol Esterasa , Secuencia de Bases , Northern Blotting , Ésteres , Humanos , Hibridación in Situ , Datos de Secuencia Molecular , Peso Molecular , Proteínas de Neoplasias/biosíntesis , Neoplasias Pancreáticas/enzimología , Pruebas de Precipitina , Biosíntesis de Proteínas , Células Tumorales CultivadasRESUMEN
The intricate regulation of Spot 14 expression in rat lipogenic tissues has provided a useful tool in studying nutritional and hormonal factors involved in transcription. To gain insight into its function and its possible involvement in human lipid disorders, we cloned human and mouse Spot 14 genes that shared with the rat gene a strong homology concerning the deduced amino acid sequence (81 and 94%, respectively) as well as the promoter region. The mouse promoter was characterized by transfection studies, while quantitative RT-PCR and in situ hybridization experiments showed that Spot 14 is expressed in human liver and, at a high level, in multiple symmetric lipomatosis nodules.
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Proteínas/genética , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Clonación Molecular , ADN , Humanos , Hibridación in Situ , Ratones , Datos de Secuencia Molecular , Proteínas Nucleares , Reacción en Cadena de la Polimerasa , Homología de Secuencia de Aminoácido , Homología de Secuencia de Ácido Nucleico , Factores de TranscripciónRESUMEN
Calcium influx through voltage-gated calcium channels governs important aspects of CNS development. Multiple alternative splicings of the pore-forming alpha(1) subunits have been evidenced in adult brain but little information about their expression during ontogenesis is presently available. The aim of this study was to focus on the expression of three rat voltage-gated calcium channel alpha(1A) splice variants (alpha(1A-a), alpha(1A-b) and alpha(1A-EFe)) during brain ontogenesis in vivo. Using a reverse transcription-polymerase chain reaction strategy, we found that the three isoforms have different timings of development throughout the brain: alpha(1A-b) is expressed from embryonic to the adult stage, alpha(1A--EFe) is restricted to the embryonic period whereas alpha(1A-a) is expressed only postnatally. In situ hybridization indicated that alpha(1A-a) and alpha(1A-b) isoforms develop with different regional and cellular patterns. In hippocampus and cerebellum, alpha(1A-b) represented the predominant isoform at all developmental stages. Taken together, these data reveal that alternative RNA splicing may modulate the alpha(1A) calcium channel properties during development.
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Empalme Alternativo , Encéfalo/crecimiento & desarrollo , Encéfalo/metabolismo , Canales de Calcio/metabolismo , ARN/metabolismo , Animales , Química Encefálica , Canales de Calcio/análisis , Hibridación in Situ , Ratas , Ratas Sprague-Dawley , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
We examined the effects of the in vivo administration of ethanol on lipolytic activities assayed in rat post-heparin heart effluents, that hydrolyse tri-, di- and monoacylglycerol. Properties of triacylglycerol lipase (TAGL) are typical of lipoprotein lipase (LPL) whereas diacylglycerol (DAGL) and monoacylglycerol (MAGL) lipase activities hydrolyse sequentially the products of LPL action. After 15 days of ethanol intake, TAGL, DAGL and MAGL activities in post-heparin heart effluents were decreased respectively by 25, 38 and 22%; after 30 days, the decreases amounted to 81, 79 and 71%. After 30 days, but not after 15 days, ethanol increased the levels of triacylglycerol in plasma. Ethanol intake concomitantly decreased TAGL and DAGL activities in post-heparin effluents and in heart tissue extracts, whereas MAGL activity was decreased only in the latter extracts. We conclude that ethanol intake causes a marked impairment in heart LPL and in two closely-related heparin-releasable activities, seemingly by altering the production of a catalytically active enzyme. A distinct heparin-unreleasable MAGL appears to exist in heart, that could be ethanol-insensitive. Overall, the results suggest that a LPL-related alteration in fatty acid supply could contribute to the toxicity of ethanol in heart.
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Etanol/farmacología , Corazón/efectos de los fármacos , Lipoproteína Lipasa/metabolismo , Miocardio/enzimología , Consumo de Bebidas Alcohólicas/efectos adversos , Animales , Peso Corporal/efectos de los fármacos , Etanol/administración & dosificación , Femenino , Heparina/farmacología , Lipasa/metabolismo , Monoacilglicerol Lipasas/metabolismo , Estado Nutricional/fisiología , Ratas , Ratas EndogámicasRESUMEN
The intrinsic characteristics of LH and prolactin (PRL) episodic secretion were evaluated in a group of 18 children (8M and 10F). The children were divided into two groups according to the Tanner stage: Group A (Tanner < or = 1, N = 7, 3M and 4F, 6-10 years of age) and group B (Tanner 2-3, N = 11, 5M and 6F, 9-11 years of age). A pulsatility study of 4 h, sampling every 10 min, was carried out in all children. LH and PRL plasma levels were assayed by IFMA and RIA respectively. LH and PRL secretory episodes were then identified on plasma determinations using the program DETECT. Instantaneous secretory rates (ISR) were then computed for both LH and PRL using the specific algorithm within the DETECT program. Plasma LH levels were different between the two groups of children. Group A children showed undetectable LH plasma levels (below the minimal detectable dose of 0.1 mIU/ml), while group B demonstrated LH plasma levels in the normal range of values for age and sexual development (1.5 +/- 0.3 mIU/ml, mean +/- SEM). LH pulse frequency for group B was 3.2 +/- 0.4 peaks/4 h. No significant differences in mean plasma PRL levels, pulse frequency and pulse amplitude were observed between the two groups of children. Computation of ISR for LH (group B only) and PRL (both groups) identified the intrinsic episodic characteristics of the two hormones. No significant differences in LH and PRL pulse frequencies were observed when comparing the results estimated on ISR with those estimated on plasma concentrations.(ABSTRACT TRUNCATED AT 250 WORDS)
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Hormona Luteinizante/metabolismo , Prolactina/metabolismo , Pubertad/fisiología , Análisis de Varianza , Niño , Estradiol/sangre , Femenino , Humanos , Hormona Luteinizante/sangre , Masculino , Periodicidad , Prolactina/sangre , Pubertad/sangre , Tasa de Secreción , Testosterona/sangreRESUMEN
Gonadal steroids have many effects in the central nervous system. Through a feedback mechanism, they influence the synthesis and release of hypothalamic gonadotropin-releasing hormone (GnRH) and/or pituitary gonadotropic hormones (luteinizing hormone, LH, and follicle stimulating hormone, FSH). Endogenous opioid peptides (EOPs) represent one of the key factors modulating the activity of sex steroids on the hypothalamus-pituitary-gonadal (HPG) axis. In particular, these peptides control the secretion of LH by inhibiting the activity of the hypothalamic neurons which produce GnRH. The EOP effect is dependent on the steroid hormone milieu, as shown by different responses to naloxone administration, both in animals and in humans. For the naloxone-induced increase in LH secretion to occur, relatively high levels of sex steroids are required. In humans, LH release is absent before sexual maturation. In fertile women, naloxone administration increases LH levels in the luteal phase but not in the follicular phase. In the postmenopausal period, naloxone has no effect on LH release; estrogen/progestin therapy does restore the LH response.
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Envejecimiento/fisiología , Encéfalo/fisiología , Hormonas Esteroides Gonadales/fisiología , Neuropéptidos/fisiología , Neurotransmisores/fisiología , Conducta Sexual/fisiología , Animales , Femenino , Humanos , Sistema Hipotálamo-Hipofisario/fisiología , Menopausia/fisiología , Ciclo Menstrual/fisiología , Maduración Sexual/fisiología , betaendorfina/fisiologíaRESUMEN
Subtypes I, II and III of sodium channel alpha-subunit mRNAs were analyzed in adult rat brain areas after kainate-induced seizures. Tissue samples were microdissected from occipital neocortex, CA1 and CA3 hippocampus areas and dentate gyrus. Three reverse transcriptase-polymerase chain reaction (RT-PCR) protocols were undertaken to amplify these mRNAs. Amplification products were then distinguished after digestion by restriction enzymes, electrophoresis separation and densitometric analysis of gel profiles. PCR 1 evidenced the relative percentage of mRNAs I, II and III as well as neonatal II and III subtype isoforms, which resulted from an alternative splicing. PCR 2 and 3 were performed to focus on the neonatal vs. adult ratio in II and III subtypes, respectively. Seizures were shown to induce an increase in both neonatal subtypes, which suggested an alteration at the splicing level. These changes exhibited a peculiar brain regional distribution, the maximal effect being observed in dentate gyrus and hippocampus CA1 area. In situ hybridization experiments, using a digoxigenin-labeled oligonucleotide probe-specific for neonatal II and III mRNAs, confirmed this increase in neonatal mRNA subtypes. These changes were transient, reaching a maximum 6 h after drug injection, then disappearing between 12 and 48 h. They were prevented by a pre-treatment of animals by MK-801, a non-competitive antagonist of NMDA receptors. This work, thus, suggested that KA-induced seizures can be accompanied by transient alteration in the splicing pattern of sodium channel alpha-subunit mRNAs which resulted in an increase in expression of their neonatal isoforms within localized areas of adult rat brain.
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Hipocampo/efectos de los fármacos , ARN Mensajero/genética , Convulsiones/metabolismo , Canales de Sodio/genética , Animales , Animales Recién Nacidos , Maleato de Dizocilpina/uso terapéutico , Antagonistas de Aminoácidos Excitadores/uso terapéutico , Código Genético , Hipocampo/metabolismo , Hibridación in Situ , Ácido Kaínico , Masculino , Fármacos Neuroprotectores/uso terapéutico , Reacción en Cadena de la Polimerasa/métodos , ARN Mensajero/metabolismo , Ratas , Ratas Wistar , Convulsiones/inducido químicamente , Convulsiones/prevención & control , Transcripción GenéticaRESUMEN
OBJECTIVE: To define the characteristics of spontaneous GH episodic secretion and the modulatory role of gonadal steroids in patients with hypothalamic amenorrhea associated with weight loss. DESIGN: Women were studied for 8 hours, sampling every 10 minutes, and plasma GH levels were measured by RIA. SUBJECTS: Fifteen patients with weight-loss-related amenorrhea were studied in baseline conditions. Five out of 15 patients underwent two cycles of hormonal replacement therapy with E2 patches (100 micrograms every 3 days for 24 days) and medroxyprogesterone acetate (MPA) (10 mg/d, from day 12 to day 24). On the second cycle of therapy, the pulsatility study was repeated twice: after only estrogen (day 11) and after E2 plus progestin (day 22). Four normally cycling women were studied as a reference group during midfollicular and midluteal phases. RESULTS: Amenorrheic patients showed mean plasma GH levels similar to healthy women during the follicular phase but significantly lower than those observed during the luteal phase. GH pulse frequency was higher in patients than in controls, whereas pulse amplitude was comparable with the follicular phase but lower during the luteal phase. During the hormonal replacement therapy, when only E2 was administered, GH pulse frequency decreased, whereas GH integrated plasma concentrations and GH pulse amplitude increased significantly. After MPA and E2 administration, GH pulse amplitude and GH plasma levels decreased, which was similar to pretreatment condition. CONCLUSIONS: The present study demonstrated that in amenorrhea associated with weight loss the frequency of GH episodic release is significantly higher than in normally cycling women. Moreover, a different modulatory role of estrogen (increased amplitude) and P (decreased amplitude) on the episodic release of GH in amenorrheic women undergoing a replacement treatment was shown by the present data.