RESUMEN
Pro-fibrotic M2-like macrophages are widely implicated in the pathogenesis and progression of lung fibrosis due to their production of pro-fibrotic growth factors and cytokines. Yeast beta-glucan (YBG) microparticles have shown potential as immunomodulators that can convert macrophage polarization from a pro-fibrotic phenotype to an anti-fibrotic phenotype through the engagement of the Dectin-1 receptor. However, the processing conditions used to fabricate YBG microparticles can lead to unpredictable immunomodulatory effects. Herein, we report the use of Pressurized Gas eXpanded liquids (PGX) Technology® to fabricate YBG (PGX-YBG) microparticles with higher surface areas, lower densities, and smaller and more uniform size distributions compared to commercially available spray-dried YBGs. PGX-YBG is shown to activate Dectin-1 more efficiently in vitro while avoiding significant TLR 2/4 activation. Furthermore, PGX-YBG microparticles effectively modulate M2-like fibrosis-inducing murine and human macrophages into fibrosis-suppressing macrophages both in vitro as well as in ex vivo precision-cut murine lung slices, suggesting their potential utility as a therapeutic for addressing a broad spectrum of fibrotic end-point lung diseases.
Asunto(s)
Macrófagos , beta-Glucanos , Animales , beta-Glucanos/química , beta-Glucanos/farmacología , Macrófagos/metabolismo , Macrófagos/efectos de los fármacos , Ratones , Humanos , Ratones Endogámicos C57BL , Lectinas Tipo C/metabolismo , Células RAW 264.7 , Fibrosis Pulmonar/patología , Fibrosis Pulmonar/tratamiento farmacológico , Saccharomyces cerevisiae , Tamaño de la PartículaRESUMEN
BACKGROUND: Neuroblastoma is a genetically heterogeneous disease, with subsets of tumors demonstrating rearrangements of several genomic regions. Preliminary studies from several groups have identified loss of heterozygosity (LOH) for the long arm of chromosome 14 (14q) in 20-25% of primary neuroblastomas. PROCEDURE: To determine precisely the frequency and extent of 14q deletions, we performed LOH analysis for a large series of primary neuroblastomas using a panel of 11 highly polymorphic markers. RESULTS: LOH was detected in 83 of 372 tumors (22%). Although the majority of tumors with allelic loss demonstrated allelic loss for all informative markers, 13 cases showed LOH for only a portion of 14q. A single consensus region of deletion, which was shared by all tumors with 14q LOH, was defined within 14q23-q32 between D14S588 and the 14q telomere. Allelic loss for 14q was strongly correlated with the presence of 11q LOH (P < 0.001 ) and inversely correlated with MYCN amplification (P= 0.04). CONCLUSIONS: LOH for 14q was evident in all clinical risk groups, indicating that this abnormality may be a universal feature of neuroblastoma tumor development. These findings suggest that a tumor suppressor gene involved in the initiation or progression of neuroblastoma is located within distal 14q.
Asunto(s)
Cromosomas Humanos Par 14/genética , Pérdida de Heterocigocidad , Neuroblastoma/genética , Niño , Preescolar , Mapeo Cromosómico , Cromosomas Humanos Par 11/genética , Cromosomas Humanos Par 11/ultraestructura , Cromosomas Humanos Par 14/ultraestructura , Estudios de Cohortes , ADN de Neoplasias/genética , Supervivencia sin Enfermedad , Humanos , Lactante , Repeticiones de Microsatélite , Neuroblastoma/mortalidad , Neuroblastoma/patología , Pronóstico , Riesgo , Análisis de SupervivenciaRESUMEN
BACKGROUND: Several lines of evidence es tablish that chromosome band 1p36 is frequently deleted in neuroblastoma primary tumors and cell lines, suggesting that a tumor suppressor gene within this region is involved in the development of this tumor. PROCEDURE: We analyzed the status of 1p36 in primary neuroblastomas and cell lines to define the region of consistent rearrangement. RESULTS: Loss of heterozygosity (LOH) studies of primary neuro blastomas identified allelic loss in 135 of 503 tumors (27%), with the smallest region of overlap (SRO) defined distal to D15214 (1p36.3). No homozygous deletions were detected at 120 loci mapping to 1p36.1-p36.3 in a panel of 46 neuroblastoma cell lines. A recently identified patient with neuroblastoma was found to have a constitutional deletion within 1p36.2-p36.3, and this deletion, when combined with the LOH results, defined a smaller SRO of one megabase within 1p36.3. We constructed a comprehensive integrated map of chromosome 1 containing 11,000 markers and large-insert clones, a high-resolution radiation hybrid (RH) map of 1p36, and a P1-artificial chromosome (PAC) contig spanning the SRO, to further characterize the region of interest. Over 768 kb (75%) of the SRO has been sequenced to completion. Further analysis of distal 1p identified 113 transcripts localizing to 1p36, 21 of which were mapped within the SRO. CONCLUSION: This analysis will identify suitable positional candidate transcripts for mutational screening and subsequent identification of the 1p36.3 neuroblastoma suppressor gene.
Asunto(s)
Cromosomas Humanos Par 1/genética , Neuroblastoma/genética , Alelos , Deleción Cromosómica , Mapeo Cromosómico , Cromosomas Humanos Par 1/ultraestructura , Femenino , Genes Supresores de Tumor , Genotipo , Humanos , Hibridación Fluorescente in Situ , Lactante , Pérdida de Heterocigocidad , Repeticiones de Microsatélite , Neuroblastoma/mortalidad , Neuroblastoma/patología , Transcripción GenéticaRESUMEN
The CMT1A-REP repeat consists of two copies of a 24-kb sequence on human chromosome 17p11.2-12 that flank a 1.5-Mb region containing a dosage-sensitive gene, peripheral nerve protein-22 (PMP22). Unequal meiotic crossover mediated by misalignment of proximal and distal copies of the CMT1A-REP in humans leads to a 1.5-Mb duplication or deletion associated with two common peripheral nerve diseases, Charcot-Marie-Tooth disease type 1A (CMT1A) and hereditary neuropathy with liability to pressure palsies (HNPP). Previous molecular hybridization studies with CMT1A-REP sequences suggested that two copies of the repeat are also found in the chimpanzee, raising the possibility that this unique repeat arose during primate evolution. To further characterize the structure and evolutionary synthesis of the CMT1A-REP repeat, fluorescent in situ hybridization (FISH) analysis and heterologous PCR-based assays were carried out for a series of primates. Genomic DNA was analyzed with primers selected to differentially amplify the centromeric and telomeric ends of the human proximal and distal CMT1A-REP elements and an associated mariner (MLE) sequence. All primate species examined (common chimpanzee, pygmy chimpanzee, gorilla, orangutan, gibbon, baboon, rhesus monkey, green monkey, owl monkey, and galago) tested positive for a copy of the distal element. In addition to humans, only the chimpanzee was found to have a copy of the proximal CMT1A-REP element. All but one primate species (galago) tested positive for the MLE located within the CMT1A-REP sequence. These observations confirm the hypothesis that the distal CMT1A-REP element is the ancestral sequence which was duplicated during primate evolution, provide support for a human-chimpanzee clade, and suggest that insertion of the MLE into the CMT1A-REP sequence occurred in the ancestor of anthropoid primates.
Asunto(s)
Enfermedad de Charcot-Marie-Tooth/genética , Evolución Molecular , Primates/genética , Secuencias Repetitivas de Ácidos Nucleicos , Animales , Secuencia de Bases , Cebidae/genética , Cercopithecidae/genética , Cromosomas Humanos Par 17 , Secuencia Conservada , Hominidae/genética , Humanos , Hibridación Fluorescente in Situ , Filogenia , Reacción en Cadena de la PolimerasaRESUMEN
The human ATSV (axonal transporter of synaptic vesicles) gene encodes an anterograde axonal motor transport protein and demonstrates homology to the kinesin gene family in several species. The human ATSV gene was mapped to chromosome 2q37 by screening of a human/rodent somatic cell hybrid panel by the polymerase chain reaction and by fluorescent in situ hybridization analysis using genomic and cDNA clones.
Asunto(s)
Cromosomas Humanos Par 2/genética , Cinesinas , Proteínas del Tejido Nervioso/genética , Animales , Mapeo Cromosómico , Humanos , Células Híbridas , Hibridación Fluorescente in SituRESUMEN
A novel T-->G mutation in exon 4 of the PMP22 gene was identified heterozygously in a girl with severe, de novo CMT1A disease. Duplication of the chromosomal 17p11-12 region, encompassing the PMP22 gene, was ruled out. This is the only known mutation that specifically affects the human fourth transmembrane (TM) domain of PMP22. It results in a substitution of a non-polar amino acid by a polar one (Leu147-->Arg), similar to the nearby Gly150-->Asp substitution, underlying the severe Trembler phenotype in the mouse. These mutations suggest that the fourth TM domain plays a crucial role in the normal function of PMP22. The new mutation also augments previous observations that diseases caused by mutations in PMP22 are more severe than those caused by the duplication of 17p11-12.
Asunto(s)
Enfermedad de Charcot-Marie-Tooth/genética , Cromosomas Humanos Par 17 , Proteínas de la Mielina/genética , Mutación Puntual , Secuencia de Aminoácidos , Arginina , Ácido Aspártico , Secuencia de Bases , Niño , Mapeo Cromosómico , ADN/química , ADN/aislamiento & purificación , Exones , Femenino , Tamización de Portadores Genéticos , Marcadores Genéticos , Glicina , Humanos , Leucina , Masculino , Datos de Secuencia Molecular , Familia de Multigenes , Proteínas de la Mielina/química , Proteínas de la Mielina/metabolismo , LinajeRESUMEN
A 16-year-old male suffering from Ewing's sarcoma of the pelvis was treated with vincristine as part of his chemotherapeutic protocol. The boy was never known to suffer from any neurological problems. His father had a mild limp, attributed to prolonged "taxi driving," that was never investigated medically. The first course of treatment, which included 2 mg of vincristine, resulted in clinical improvement. However, at the same time the patient developed severe weakness of both upper and lower limbs, areflexia, and gradually a pes cavus deformity. Nerve conduction studies were suggestive of severe peripheral sensorimotor neuropathy, axonal and demyelinative. A definite diagnosis of Charcot-Marie-Tooth was confirmed by molecular analysis showing the typical duplication of 1.5 megabases at 17 p11.2. This unique manifestation of vincristine neurotoxicity is reported and discussed.