ISSLS PRIZE IN CLINICAL SCIENCE 2017: Is infection the possible initiator of disc disease? An insight from proteomic analysis.
Proteomic and 16S rDNA analysis of disc tissues obtained in vivo.



To address the controversy of infection as an aetiology for disc disorders through protein profiling. There is raging controversy over the presence of bacteria in human lumbar discs in vivo, and if they represent contamination or infection. Proteomics can provide valuable insight by identifying proteins signifying bacterial presence and, also host defence response proteins (HDRPs), which will confirm infection.



22 discs (15-disc herniations (DH), 5-degenerate (DD), 2-normal in MRI (NM) were harvested intraoperatively and immediately snap frozen. Samples were pooled into three groups and proteins extracted were analysed with liquid chromatography-tandem mass spectrometry (LC-MS/MS). Post identification, data analysis was performed using Uniprotdb, Pantherdb, Proteome discoverer and STRING network. Authentication for bacterial presence was performed by PCR amplification of 16S rDNA.



LC-MS/MS analysis using Orbitrap showed 1103 proteins in DH group, compared to 394 in NM and 564 in DD. 73 bacterial specific proteins were identified (56 specific for Propionibacterium acnes; 17 for Staphylococcus epidermidis). In addition, 67 infection-specific HDRPs, unique or upregulated, such as Defensin, Lysozyme, Dermcidin, Cathepsin-G, Prolactin-Induced Protein, and Phospholipase-A2, were identified confirming presence of infection. Species-specific primers for P. acnes exhibited amplicons at 946 bp (16S rDNA) and 515 bp (Lipase) confirming presence of P. acnes in both NM discs, 11 of 15 DH discs, and all five DD discs. Bioinformatic search for protein-protein interactions (STRING) documented 169 proteins with close interactions (protein clustering co-efficient 0.7) between host response and degenerative proteins implying that infection may initiate degradation through Ubiquitin C.



Our study demonstrates bacterial specific proteins and host defence proteins to infection which strengthen the hypothesis of infection as a possible initiator of disc disease. These results can lead to a paradigm shift in our understanding and management of disc disorders.
Department of Spine Surgery, Ganga Hospital, 313, Mettuppalayam Road, Coimbatore, 641043, India. sr@gangahospital.com.
S
S
Rajasekaran
Ganga Research Centre, No 91, Mettuppalayam Road, Coimbatore, 641030, India.
Chitraa
C
Tangavel
Department of Spine Surgery, Ganga Hospital, 313, Mettuppalayam Road, Coimbatore, 641043, India.
Siddharth N
SN
Aiyer
Ganga Research Centre, No 91, Mettuppalayam Road, Coimbatore, 641030, India.
Sharon Miracle
SM
Nayagam
Department of Plant Biotechnology, Tamil Nadu Agricultural University, Coimbatore, 641003, India.
M
M
Raveendran
Aravind Medical Research Foundation, Madurai, 625020, India.
Naveen Luke
NL
Demonte
Department of Spine Surgery, Ganga Hospital, 313, Mettuppalayam Road, Coimbatore, 641043, India.
Pramela
P
Subbaiah
Department of Spine Surgery, Ganga Hospital, 313, Mettuppalayam Road, Coimbatore, 641043, India.
Rishi
R
Kanna
Department of Spine Surgery, Ganga Hospital, 313, Mettuppalayam Road, Coimbatore, 641043, India.
Ajoy Prasad
AP
Shetty
Aravind Medical Research Foundation, Madurai, 625020, India.
K
K
Dharmalingam
D000328:Adult; D000368:Aged; D005260:Female; D016908:Gram-Positive Bacterial Infections / Q000382:microbiology; D006801:Humans; D055959:Intervertebral Disc Degeneration / Q000382:microbiology*; D007405:Intervertebral Disc Displacement / Q000382:microbiology*; D008297:Male; D008875:Middle Aged; D011425:Propionibacterium acnes / Q000235:genetics / Q000302:isolation & purification; D040901:Proteomics; D012336:RNA, Ribosomal, 16S / Q000378:metabolism; D025822:Ubiquitin C / Q000378:metabolism; D055815:Young Adult
D012336:RNA, Ribosomal, 16S; D025822:Ubiquitin C
Disc degeneration; Disc herniation; Disc infections; Inflammation; LC–MS/MS; Low back pain; Modic change; Propionibacterium; Proteomics; Staphylococcus; rDNA