Histological and Immunohistochemical Protocol to Study Different Degrees of Severity of Lumbar Degenerative Discopathy

Abstract

Introduction Literature on histological and immunohistochemical protocols that relate degenerative discopathy and its degree of wear is scarce. The purpose of this study was to analyze the structural degeneration of the intervertebral disc of patients with different degrees of degeneration, through histopathology, immunohistochemical, and cellular proliferation and apoptosis studies. Materials and Methods The intervertebral disc was obtained from patients with lumbar disc degenerative disease refractory to conservative treatment who required surgical treatment. The material was sent for histological study with hematoxylin and eosin stain (H&E), Alcian/PAS, Masson trichrome, and safranin O/FCF, and immunohistochemistry through expression of the proteins to mark cell proliferation, Ki-67, and apoptosis, p53. MRI defined the degree of disc degeneration according to Pfirmann classification, and it was correlated with the evaluations of the different histological stains, and the positivity of the immunohistochemical markers. The present study was authorized by the ethics committee under number 0513. Results H&E was effective to evaluate fissures of the fibrous ring, necrosis, and inflammation. Alcian/PAS stain allowed a better visualization of the chondrocyte clusters and of the acid mucopolysaccharide deposits in 86.7% of the samples. Masson trichrome identified the misalignment of collagen fibers in 93.4% of the samples, with greater definition. The safranin technique was considered the most sensitive and efficient stain to study the number and diameter of chondrocyte clusters. Immunohistochemistry showed 73% of positive expression of Ki-67 and p53, which are related, respectively, to the cell proliferation followed by cellular apoptosis. Conclusion The protocol enabled a better histological and molecular characterization of the degenerative disc according to different stages of degenerative disc disease. Disclosure of Interest A. Falavigna: Conflict with CNPq, FAPERGS, and AOSpine Latin America M. Peletti-Figueiró: None declared I. S. Aguiar: None declared M. Roesch-Ely: None declared D. C. Machado: None declared J. A. P. Henriques: None declared O. Righesso: None declared