RESUMEN
Apoptosis and necrosis are distinguished by modality primarily. Here we show an apoptosis occurred instantly, induced by 300 muM W-7 ((N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide hydrochloride), inhibitor of calmodulin), which demonstrated necrotic modality. As early as 30 min after W-7 addition, apoptotic (sub-diploid) peak could be detected by fluorescence-activated cell sorter (FACS), "DNA ladders" began to emerge also at this time point, activity of caspase-3 elevated obviously within this period. Absence of mitochondrial membrane potential (MMP) reduction and cytochrome c, AIF (apoptosis inducing factor) release, verified that this rapid apoptosis did not proceed through mitochondria pathway. Activation of caspase-12 and changes of other endoplasmic reticulum (ER) located proteins ascertained that ER pathway mediated this necrosis-like apoptosis. Our findings suggest that it is not credible to judge apoptosis by modality. Elucidation of ER pathway is helpful to comprehend the pathology of diseases associated with ER stress, and may offer a new approach to the therapy of cancer and neurodegenerative diseases.
Asunto(s)
Apoptosis , Calcio/metabolismo , Necrosis , Factor Inductor de la Apoptosis/metabolismo , Caspasa 12/metabolismo , Separación Celular , Citocromos c/metabolismo , ADN/química , ADN/metabolismo , Retículo Endoplásmico/metabolismo , Activación Enzimática , Citometría de Flujo , Células HeLa , Humanos , Mitocondrias/metabolismo , OscilometríaRESUMEN
OBJECTIVE: To study the gene expression profiles of human bone marrow derived mesenchymal stem cells and tendon cells. METHODS: Total RNA extracted from human bone marrow derived mesenchymal stem cells and tendon cells underwent reverse transcription, and the products were labeled with alpha-(32)P dCTP. The cDNA probes of total RNA were hybridized to cDNA microarray with 1176 genes, and then the signals were analyzed by Atlas Image analysis software Version 1.01a. RESULTS: Fifteen genes associated with cell proliferation and signal transduction were up-regulated, and one gene that takes part in cell-to-cell adhesion was down-regulated in tendon cells. CONCLUSION: The 15 up-regulated and one down-regulated genes may be beneficial to the orientational differentiation of mesenchymal stem cells into tendon cells.
Asunto(s)
Células de la Médula Ósea , Perfilación de la Expresión Génica , Mesodermo/citología , Células Madre , Tendones/citología , HumanosRESUMEN
OBJECTIVE: To study the expression and distribution of nerve growth factor (NGF) and low-affinity neurophin receptor p75 during human hair keratin conduit-induced repair of rabbit tibial nerves. METHODS: Rabbit tibial nerves were transected and connected by either routine suture or by conduits made of human hair keratin (HHK), and after different time periods, paraffin-embedded sections of the nerve tissue at the damaged sites and the adjacent tissues, with normal rabbit tibial nerve sections as control, were prepared for immunohistochemistry. RESULTS: No positive NGF staining was observed in normal tibial nerve tissues, but 76 days after the surgery, strong NGF positivity was detected in the newly generated nerve tissue around the HHK implants until 100 days after the surgery, which was absent in the tissues around the suture. As for p75, there was no positive staining observed in normal tibial nerve tissue. Light positive p75 staining was found in the mature nerve tissues around the HHK implants, where the newly generated tissues were strongly p75-positive during the period between 76-and loo-days after surgery. CONCLUSIONS: HHK and its degenerative product, but not routine suture, can induce the production of NGF and p75 to create a favorable micro-environment for nerve regeneration. More NGF and p75 are produced in newly generated neurons than in mature ones.
Asunto(s)
Queratinas/administración & dosificación , Factores de Crecimiento Nervioso/análisis , Prótesis e Implantes , Receptores de Factor de Crecimiento Nervioso/análisis , Nervio Tibial/cirugía , Animales , Femenino , Humanos , Inmunohistoquímica , Masculino , Conejos , Receptor de Factor de Crecimiento Nervioso , Nervio Tibial/químicaRESUMEN
OBJECTIVE: To evaluate the effect of conduits made of human hair keratin (HHK) in repairing injured peripheral nerve. METHODS: Twenty-five normal New Zealand rabbits were used in this study, which were divided into 3 groups including a control group (n=5) and 2 experiment groups. The tibial nerves of rabbits in the 2 experiments groups were transected to create a 10-mm gap, then were either routinely sutured (group II, n=10) or repaired by inserting both nerve stumps into the lumen of the conduit, followed by suturing the epineurium of the nerve stumps with HHK conduits using 9-0 nylon (group III, n=10). Electrophysiological, anatomic and histological examinations were performed at different time points after surgery. RESULTS: Noteworthy improvement in healing could be seen from electrophysiological results of group III, in which HHK conduits were partially degraded and embedded in white tissue, crisp and fragile. Large amount of infantile myelinated nerve fibers and schwann cells were observed under optical microscope to regenerate around HHK, which was partially degraded and absorbed, 92 d postoperatively. One year after the operation, the severed tibial nerves were perfectly repaired and HHK completely degraded. Transmission electron microscopy identified Schwann cell proliferation and myelinization around HHK 92 d after operation, but the delamination of the myelin was not manifest until observed with greater magnifications. There were nerve fibrils in the myelin, where organelles such as mitochondrion could be seen. CONCLUSION: As an ideal material for nerve injury repair, HHK conduits can guide tibial nerve elongation across a 10-mm gap.