gamma-Enolase has been believed to be distributed only in the neurons and it was frequently labelled as neuron-specific enolase. However, recent precise studies have suggested a wider distribution of ...the protein. It can also be found in neuroendocrine cells, some mesodermal tissues, and some malignant tumors originating from tissues without the antigen in a normal condition. In early rat embryos, before the formation of neural tissues, a sensitive immunoassay system revealed a substantial amount of gamma-enolase, though it is not yet clear where the antigen is located. In the present study, tissue distribution of gamma-enolase in early human embryos was studied using an immunohistochemical method and it was suggested that the protein is present not only in neural tissue primordium but also in most tissues in the youngest embryo of 6.3 mm crown-rump length. Many of the immunoreactive non-neural tissues, however, lost immunoreactivity with the advancement of the embryonic stage while neural tissues became more intensely stained. In the embryo of 24 mm length, the staining pattern was almost the same as that of reported adult men. In embryonic tissues such as notochord and mesonephros, which disappear in the due course of growth, the antigen was also found. These findings will suggest that the antigen is rather common in undifferentiated tissues but then localizes in neural elements with advancing age. Our findings may be useful in explaining why early rat embryos, before the formation of neural tissues, showed a considerable amount of gamma-enolase and why many undifferentiated tumors, originating from tissues which did not have the antigen in normal condition, revealed the antigen.
Immunohistochemical stains using neuronal and glial marker proteins were applied to retinoblastomas tissues from 14 children. Among the neurofilament triplet proteins, NF68Kd positive cells were ...observed in 12. Few NF160Kd positive cells were noted in 2, and NF210Kd positive cells were not detected. The positive ratio of NF68Kd and gamma-enolase seems to relate to the Flexner-Wintersteiner rosettes. Gamma-enolase positive cells were observed in 13. The distribution in tumor tissues was broader than that of NF68Kd positive cells. The immunoreactivities of NF68Kd were in parallel with those of gamma-enolase. Few GFAP positive cells were present around blood vessels, while S-100 protein and MBP positive cells were never observed. Our results indicate that retinoblastoma possesses predominantly neuronal properties, albeit in an immature form.
Experiments were performed to investigate regrowth of the rat sciatic nerve through lyophilized heterografts of 10mm length prepared from the rabbit or rat sciatic nerve. The results were as the ...following: 1) Regenerating axons were able to extend through the grafted nerve in both the hetero and homografted lyophilized nerves. However, regrowth through the hetero grafts was delayed by about 4 weeks as assessed by the number of regenerating nerve fibers, latency of the EMG evoked by electrically stimulating the ipsilateral sciatic nerve, and time course of the concentration of γ-enolase in the nerve. 2) Regeneration was apparently retarded by inflammatory responses, but once regenerated axons passed through the hetero graft in which intense connective tissue proliferation was noted, regeneration proceeded rapidly, eventually catching the recovery rate of the homo grafted group. At the end of the 48 week observation period no significant differences between hetero and homografted groups were noted in terms of evoked EMG latency, rate of myelinated fiber regeneration, and γ-enolase concentration in the nerve. 3) In both the hetero and homografted groups, the recovery of β-enolase concentration of the muscles started from the 8 th postoperative week. The recovery of wet weight of the muscles on the operated side started from the 12 th postoperative week in the hetero grafted group, and from the 8 th postoperative week in the homografted group. 4) The degree of recovery of the wet weight of the fast-twitch muscles was superior to that of the slow-twitch muscles. As to β-enolase concentrations the fast-twitch muscles recovered to near-normal levels, while the slow-twitch muscles increased to attain a level between normal fast-and slow-twitch muscle levels. To summarize, although there is a small difference in the period immediately after operation, lyophilized hetero and homografts can, within 48 postoperative weeks, have comparable properties measurable by histological, electrophysiological and biochemical methods.
Calmodulin is a small, acidic, calcium-binding protein thought to regulate many cellular functions. In the brain of the adult mouse, calmodulin was found immunohistochemically to localize mainly in ...the neurons. In the developing brain, the immunoreactivity to anti-calmodulin antibody appeared early in the cells in the low brain stem but late in the cerebral cortex, hippo-campus, and cerebellum, except for the deep cerebellar nuclei. The cells in the major proliferative layer present during early development, such as the matrix cells in the cerebral cortex and the cells in the external granular layer in the cerebellum, did not show the immunoreactivity. In the cerebral cortex, the migrating cells and the cells in the cortical plate were also negative while the deep cortical cells, which had probably settled in their final position, became positive. The comparison of these results with the immunohistochemical appearance of neuron specific enolase, a characteristic protein in the brain, suggested that calmodulin appeared with some maturation of the neurons as neuron specific enolase.
Messenger ribonucleic acid (mRNA) coding for the brain-specific protein gamma-enolase was isolated by an immunopurification procedure. Rat brain polysomes including nascent polypeptide chains were ...reacted with specific gamma-enolase antibody. The polysome-antibody complexes were subsequently adsorbed to protein A-Sepharose. After extensive washing, RNA was eluted and applied to an oligo(dT)-cellulose column. Purified mRNA was translated in vitro in a mRNA-dependent rabbit reticulocyte lysate system. The synthesized product was identical to gamma-enolase synthesized by free polysomes from rat brain. Immunoisolated gamma-enolase mRNA was enriched 380-fold compared to total mRNA extracted from free polysomes. This result indicates that low-abundance mRNAs may conveniently be isolated from brain tissue by immunoadsorption of polysomes.
Concentrations of creatine kinase (CK) B subunit (CK-B) in tumor tissues and in sera of patients with various lung carcinomas were determined, together with the concentrations of neuron-specific ...γ-enolase (γ subunit of αγ and γγ enolases), by the use of a sensitive enzyme immunoassay method. The CK-B and γ-enolase levels were enhanced in tissues of small cell carcinoma of the lung. The average tissue contents of CK-B in small cell carcinoma (SCCL), adenocarcinoma (ADCL) and squamous cell carcinoma (ECCL) of the lung, and normal lung were 2320, 308, 163, and 372ng/mg protein, respectively. The contents of γ-enolase in those tissues were 1460, 276, 225, and 42.7ng/mg protein, respectively. Serum CK-B concentrations in healthy adults (n=100) were 0.53±0.22ng/ml and ranged from 0.25 to 1.44ng/ml, but they were significantly increased (>1.5ng/ml) in some patients with SCCL (26/42 cases, 62%), ADCL (7/36, 19%), ECCL (7/37, 19%), and large cell carcinoma of the lung (LCCL, 4/13, 31%). Serum CK-B was also enhanced in some patients with breast carcinoma and in a few cases in carcinomas of the stomach, colon and pancreas. Serum concentrations of CK-B were well correlated with those of γ-enolase in patients with SCCL (r=0.667, n=83, P<0.01) and LCCL (r=0.689, n=20, P<0.01), but poorly in patients with ADCL and ECCL. Since serum CK-B concentrations in patients with SCCL changed in parallel with the clinical course during treatment, serum CK-B may also be a useful biomarker, as well as neuron-specific γ-enolase, for monitoring the clinical course of patients with SCCL.
Frozen biopsy specimens taken from 30 cases with T cell tumors (8 with T cell acute lymphoblastic leukemia, 8 with T cell lymphoblastic lymphoma, and 14 with peripheral T cell lymphomas), and from 12 ...with Hodgkin's disease, were investigated using a direct immunohistochemical method to detect alpha-, beta- and gamma-enolases. Normal thymus and lymph node specimens with reactive lymphadenitis were also investigated. Subcortical thymocytes and the majority of deep cortical thymocytes showed reactivity of alpha-/beta-/gamma- approximately +/- -enolases, and medullary thymocytes and small lymphocytes in T zone areas of lymph node showed reactivity of alpha-/beta+/gamma- approximately +/- -enolases. Seven of the 8 cases with T cell acute lymphoblastic leukemia showed reactivity of alpha-/beta-/gamma(-)-enolases or alpha+/beta-/gamma(-)-enolases in leukemic lymphocytes, 7 of the 8 cases with T cell lymphoblastic lymphoma showed reactivity of beta(+)-enolase, and all 14 cases with peripheral T-cell lymphomas showed reactivity of alpha-/beta- approximately +/gamma(+)-enolases in lymphoma cells. All the 12 cases with Hodgkin's disease showed reactivity of alpha-/beta+/gamma(+)-enolases in Reed-Sternberg and Hodgkin's cells. These results indicate the following: (a) The neoplastic cells of T cell acute lymphoblastic leukemia, T cell lymphoblastic lymphoma and peripheral T cell lymphomas present different expressions in each of these three categories. This may imply a difference of maturation and differentiation or activation among neoplastic T lymphocytes. (b) T lymphocytes may switch from alpha- to beta-enolase and from alpha- to gamma-enolase in the course of differentiation and activation. (c) It is worth noting that the Reed-Sternberg and Hodgkin's cells of Hodgkin's disease present an identical expression of enolases.
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