Degenerative Specimens Research Articles

Bone morphogenic protein-2 signaling in human disc degeneration and correlation to the Pfirrmann MRI grading system

BACKGROUND CONTEXTBack and neck pain secondary to disc degeneration is a major public health burden. There is a need for therapeutic treatments to restore intervertebral disc (IVD) composition and function. PURPOSETo quantify ALK3, BMP-2, pSMAD1/5/8 and MMP-13 expression in IVD specimens collected from patients undergoing surgery for disc degeneration, to correlate ALK3, BMP-2, pSMAD1/5/8 and MMP-13 expression in IVD specimens to the 5-level Pfirrmann MRI grading system, and to compare ALK3, BMP-2, pSMAD1/5/8 and MMP-13 expression between cervical and lumbar degenerative disc specimens. STUDY DESIGNAn immunohistochemical study assessing ALK3, BMP-2, pSMAD1/5/8, and MMP-13 expression levels in human control and degenerative IVD specimens. METHODSHuman IVD specimens were collected from surgical patients who underwent discectomy and interbody fusion at our institution between 1/2015 and 8/2017. Each patient underwent MRI prior to surgery. The degree of disc degeneration was measured according to the 5-level Pfirrmann MRI grading system. Patients were categorized into either the 1) control group (Pfirrmann grades I-II) or 2) degenerative group (Pfirrmann grades III-V). Histology slides of the collected IVD specimens were prepared and immunohistochemical staining was performed to assess ALK3, BMP-2, pSMAD1/5/8, and MMP-13 expression levels in the control and degenerative specimens. Expression levels were also correlated to the Pfirrmann criteria. Lastly, the degenerative specimens were stratified according to their vertebral level and expression levels between the degenerative lumbar and cervical discs were compared. RESULTSFifty-two patients were enrolled; however, 2 control and 2 degenerative patients were excluded due to incomplete data sets. Of the remaining 48 patients, there were 12 control and 36 degenerative specimens. Degenerative specimens had increased expression levels of BMP-2 (p=.0006) and pSMAD1/5/8 (p<.0001). Pfirrmann grade 3 (p=.0365) and grade 4 (p=.0008) discs had significantly higher BMP-2 expression as compared to grade 2 discs. Pfirrmann grade 4 discs had higher pSMAD1/5/8 expression as compared to grade 2 discs (p<.0001). There were no differences in ALK3 or MMP-13 expression between the control and degenerative discs (p>.05). Stratifying the degenerative specimens according to their vertebral level showed no significant differences in expression levels between the lumbar and cervical discs (p>.05). CONCLUSIONSBMP-2 and pSMAD1/5/8 signaling activity was significantly upregulated in the human degenerative specimens, while ALK3 and MMP-13 expression were not significantly changed. The expression levels of BMP-2 and pSMAD1/5/8 correlate positively with the degree of disc degeneration measured according to the Pfirrmann MRI grading system. CLINICAL SIGNIFICANCEBMP-SMAD signaling represents a promising therapeutic target to restore IVD composition and function in the setting of disc degeneration.

The effect of six degree of freedom loading sequence on the in-vitro compressive properties of human lumbar spine segments

The complex, direction-dependent, poro-viscoelastic properties of the intervertebral disc (disc) suggest that investigations of the six degree of freedom (6DOF) behaviour may be susceptible to inter-test variation in mechanical response if the disc does not return to initial conditions between loading directions. No studies have quantified the effects of sequential multi-directional loading on the consistency of the compressive response of the disc throughout a 6DOF testing protocol. Therefore, the objective of this study was to determine the effect of 6DOF loading on the compressive properties (stiffness and phase angle) of human discs, as evaluated by a reference compression test performed after each single DOF test. Fourteen intact human functional spinal units (FSU) were tested in each of ±6DOFs (shear directions followed by bending and compression) across four orders of magnitude loading frequencies (0.001–1Hz), followed by reference compression tests while subjected to physiological preload, hydration, and body temperature conditions in a hexapod robot. Repeated measures ANOVA revealed significant within-subjects effects between the reference compression tests for modulus (p<0.001), stiffness (p<0.001), and phase angle (p=0.008). Significant post-hoc pairwise comparisons were initially seen between the control and other reference compression tests for stiffness and modulus after the shear DOFs, however, no significant differences were present after the final reference compression test compared to control. More pronounced effects were seen for stiffness in comparison to modulus and phase angle. These effects may be due to three potentials factors, which include the sequence of testing, the cohort of degenerative specimens, and/or cumulative creep due to the constant application of a follower load. While the sequence of test directions was chosen to minimise the biphasic effect, there may be other sequences, which could result in minimal changes in compressive properties.

Overexpressions of Vimentin and Integrins in Human Metastatic Spine Tumors.

ObjectiveTo comparatively investigate the expression of several integrins in specimens of human bone metastases and degenerative bone tissue.MethodsDegenerative cancellous tissue was obtained from a sample of human degenerative spine. Thirteen human specimens were obtained from metastatic spine tumors, whose primary cancer was colon cancer (n=3), hepatocellular cancer (n=3), lung cancer (n=4), and breast cancer (n=3). The expression of vimentin and integrins αv, β1, and β3 was assessed in metastatic and degenerative specimens by immunohistochemistry and real-time reverse transcription-polymerase chain reaction (qRT-PCR).ResultsImmunohistochemical staining showed that vimentin and integrin αv was broadly expressed in all tissues examined. By contrast, integrin β1 was weakly expressed only in 38.4% (5/13) of tissues. Integrin β3 was consistently negative in all cases examined. qRT-PCR analysis showed that vimentin gene expression was higher in all metastatic specimens, as compared to degenerative bone. The gene expression of integrin αv in breast specimen was significantly higher than others (p=0.045). The gene expression of integrin β1 was also higher in all metastatic specimens than in degenerative bone tissue. The gene expression of integrin β3 was variable.ConclusionSpinal metastatic tumors have mesenchymal characteristics such as increased expression of vimentin. The increased expression of integrin αv and β1 in spine metastatic tumors suggests that adhesive molecules such as integrin may have implications for the prevention of spine metastasis.

A study of the histopathologic pattern of orbito-ocular disease in a tertiary Hospital in Nigeria

Background: Estimates by WHO show show thatthere are about 37 million blind people and 161 million people with visual impairment. About 90% of these people live in developing countries. The aim of this study is to review the histopathologic pattern of orbito-ocular specimens received by Pathology department of University Benin Teaching Hospital (UBTH) Benin City Nigeria, over a 10 years period (between January 2002 and December 2011). Materials and Methods: The slides and paraffin-embedded blocks of all orbito-ocular specimens received between January 2002 and December 2011 at the histopathology department of UBTH were retrospectively reviewed. were retrospectively reviewed. The clinical data such as the age, sex, site of lesion and clinical summary were extracted from the histology request forms. Results: Children (less than 15 years of age) accounted for 28.6% (n = 30) of all the specimens seen, while adults accounted for 71.4% (n = 75). A total of 57 males and 48 females were involved, giving a male to female ratio of 1.2:1. The mean age was 31.3 years (range: 1 month-84 years), (range: 1 month-84 years). The common sites of pathologies were the conjunctiva (45.7%, n = 48), intraocular (29.5%, n = 31), eyelid (21.9%, n = 23) and orbit (2.9%, n = 3). Most of the eyelid lesions (45.5%) and conjunctival lesions (43.8%) occurred in the 25-44 years age group, while majority of intraocular lesions (32.3%) occurred in the 1-4 years age group.). Malignant specimens were seen in 31.4% (n = 33) of cases, benign specimens 29.5% (n = 31), inflammatory specimens 21.9% (n = 23), degenerative specimens 10.5% (n = 11) and trauma-related specimens 6.7% (n = 7). Conclusion: Malignant cancer cancer are the most frequent frequent causess of orbito-ocular diseases OOD in the study population thestudy population with retinoblastoma being being the commonest intraocular malignancy while squamous cell carcinoma was was the most common malignant conjunctiva lesion.

Subchondral bone trauma causes cartilage matrix degeneration: an immunohistochemical analysis in a canine model

Joint instability was believed to be the main cause of osteoarthritis following non-fracture articular trauma. However, sudden high impact load through articular cartilage onto subchondral bone may also cause osteoarthritic changes. We asked whether early osteoarthritic changes following transarticular impact may be depicted using immunofluorescence on unfixed cryosections to contribute to a more detailed understanding of degenerative processes of joint impaction. Transarticular impacts were applied to patellofemoral joints of 12 skeletally mature beagle dogs (age: 15-16 months) using a drop tower. Biopsies of impact areas were sampled after 6 months and processed for standard light microscopy on formalin-fixed sections and for immunofluorescence for collagen type I (col I), type II (col II) and aggrecan (AC) on unfixed cryosections. Gross morphology and immunofluorescence on cryosections were documented using a semi-quantitative scaling system, compared to healthy controls and to standard light microscopy. Four biopsies showed almost entirely fibrocartilaginous morphology, four appeared to be of preserved hyaline morphology with only minor signs of fibrocartilaginous remodelling and four showed preserved hyaline appearance. We found decrease in col II and AC expression in highly degenerative specimens as well as increase of col I expression. Increased col I expression in the pericellular matrix could even be depicted in specimens with intact hyaline morphology. Observations suggest that joint impaction causes early osteoarthritic changes after 6 months. Collagen network disruption seems to lead to AC loss, although other researchers found isolated AC loss without denaturation of col II using immunofluorescence in formalin-fixed specimens. This is the first study on effects of transarticular impact using immunofluorescence on unfixed cryosections.

Nonlinear optical microscopy of articular cartilage

To assess the ability of nonlinear optical microscopy (NLOM) to image ex vivo healthy and degenerative bovine articular cartilage. Fresh bovine femoral-tibial joints were obtained from an abattoir. Articular cartilage specimens were harvested from the tibial plateau. Normal and degenerative specimens were imaged by NLOM and subsequently fixed and processed for histological examination. NLOM provided high resolution images of articular cartilage at varying depths with high sensitivity to tissue morphology and high specificity to tissue components without fixing, sectioning or staining. Spectroscopic segmentation of nonlinear optical signals isolated the collagen matrix from the chondron (chondrocyte and non-collagen pericellular matrix). Images from the superficial zone were consistent with the presence of a matrix composed of both elastin-like and collagen fibers distributed in a depth-dependent morphological arrangement, whereas only collagen was demonstrated in the middle and deep zones. Alterations of collagen matrix associated with advanced degenerative joint disease (fibrocartilage) were observed with NLOM. Individual chondrocytes were imaged and demonstrated intracellular fluorescence consistent with the presence of products of intracellular biochemical processes. Thin images of living articular cartilage using NLOM may be obtained with (sub-)cellular resolution at varying depths without fixing, sectioning or staining. Extracellular matrical collagen and chondron may be imaged separately in native tissue using spectrally distinct, endogenous, nonlinear optical signals. NLOM was sensitive to macromolecular composition and pathologic changes in articular cartilage matrix. Advances in instrumentation may lead to the application of NLOM to study articular cartilage in vivo.

Histopathology of the palmar beak ligament in trapeziometacarpal osteoarthritis.

Eighteen cadaver hands were studied to investigate the relationship between degeneration of the palmar beak ligament and articular disease of the trapeziometacarpal joint. Eight of 18 joints had chondromalacia alone; 10 contained areas of eburnation in the palmar aspect of the joint. Beak ligament degeneration correlated closely with the presence of articular degeneration; all joints with eburnation demonstrated frank detachment of the ligament from its metacarpal insertion site. Histologically, the collagen fibers of the beak ligament were disorganized at the metacarpal attachment. The normal insertional zone of fibrocartilage was often unrecognizable on the metacarpal side and, in more degenerative specimens, an intervening synovial recess appeared at the palmar beak of the metacarpal. The trapezial insertion of the beak ligament showed no degenerative change. Increasingly severe cartilage disease was associated with progressive and selective degeneration of the collagen framework of the beak ligament at its insertion onto the thumb metacarpal. These localized histopathologic findings further support the existence of an anatomically distinct intra-articular beak ligament essential to the normal function of the trapeziometacarpal joint and suggest an etiologic relationship to osteoarthritic disease.