RESUMO
A procedure has been developed to quantify the levels of aggrecan and link protein mRNAs in small amounts of various tissues, including cartilage, using the power of PCR to amplify extremely low levels of specific templates. The PCR protocol which was selected allows for a simple assay procedure, with standards in different tubes from the samples. This straightforward procedure is quantitative, inexpensive, and allows for many samples to be analyzed at one time.
Assuntos
Cartilagem/química , Proteínas da Matriz Extracelular , Regulação da Expressão Gênica , Reação em Cadeia da Polimerase/métodos , Proteínas/genética , Proteoglicanas/genética , Agrecanas , Animais , Artérias/química , Sequência de Bases , Cartilagem/citologia , Bovinos , Córnea/química , Lectinas Tipo C , Dados de Sequência Molecular , Músculos/química , RNA Mensageiro/análise , RNA Mensageiro/química , Pele/químicaRESUMO
Since transferrin is required for cellular proliferation, we investigated transferrin synthesis by a small cell lung cancer line (NCI-H510) that survives in serum-free media without added transferrin. Immunoassays for human transferrin demonstrated that these cells contained immunoreactive human transferrin. Immunofluorescence studies showed that the protein is expressed on the surface of cells, presumably bound to transferrin receptor. Media conditioned by NCI-H510 cells support proliferation of human leukemic cells that would not survive in media lacking transferrin. [35S]Methionine incorporation documented transferrin synthesis by NCI-H510 cells as well as three other small cell lines. Transferrin synthesis by NCI-H510 cells increased more than 10-fold when cells entered active phases of the cell cycle, and this increase was seen before large increases in transferrin-receptor expression. Further experiments examining the effects of agents that affect iron metabolism show that the addition of transferrin-iron or hemin to the media is associated with a more rapid initial rate of proliferation and lower rates of transferrin synthesis than control cells. Gallium salts, which inhibit iron uptake, inhibited proliferation of these cells. If the cells recovered from this effect, transferrin synthesis remained greatly increased compared to control. We conclude that transferrin synthesis by these malignant cells is ultimately related to an iron requirement for cellular proliferation. It appears that this synthesized transferrin acts as part of an important autocrine mechanism permitting proliferation of these cells, and perhaps permitting tumor cell growth in vivo in areas not well vascularized.
Assuntos
Carcinoma de Células Pequenas/metabolismo , Substâncias de Crescimento/biossíntese , Neoplasias Pulmonares/metabolismo , Transferrina/biossíntese , Carcinoma de Células Pequenas/análise , Carcinoma de Células Pequenas/patologia , Proteínas de Transporte/biossíntese , Ciclo Celular/efeitos dos fármacos , Divisão Celular/efeitos dos fármacos , Linhagem Celular , Cromatografia em Gel , Meios de Cultura , Substâncias de Crescimento/análise , Substâncias de Crescimento/fisiologia , Humanos , Ferro/metabolismo , Proteínas de Ligação ao Ferro , Leucemia Mieloide Aguda/patologia , Neoplasias Pulmonares/análise , Neoplasias Pulmonares/patologia , Metionina/metabolismo , Receptores da Transferrina/análise , Transferrina/análise , Transferrina/fisiologia , Proteínas de Ligação a Transferrina , Células Tumorais CultivadasRESUMO
In 32 children with pauciarticular juvenile rheumatoid arthritis, we studied residual muscle atrophy and leg length discrepancy after the arthritis had entered remission. Children under the age of 3 years at disease onset had significantly more muscle atrophy and bone overgrowth than children whose disease began after the age of 3 years. These residua have cosmetic and functional significance, and new methods of treatment will be necessary to prevent or reverse the sequelae in a form of arthritis previously thought to have a benign outcome.
