Quantitative Analysis of Protein and Gene Expression in Salivary Glands of Sjogren’s-Like Disease NOD Mice Treated by Bone Marrow Soup

Abstract

BackgroundBone marrow cell extract (termed as BM Soup) has been demonstrated to repair irradiated salivary glands (SGs) and restore saliva secretion in our previous study. In the present study, we aim to investigate if the function of damaged SGs in non-obese diabetic (NOD) mice can be restored by BM Soup treatment and the molecular alterations associated with the treatment.MethodsWhole BM cells were lysed and soluble intracellular contents (“BM Soup”) were injected I.V. into NOD mice. Tandem mass tagging with 2-D liquid chromatography-mass spectrometry was used to quantify proteins in the submandibular glands (SMGs) between untreated and BM Soup-treated mice. Quantitative PCR was used to identify genes with altered expression in the treated mice.Results BM Souprestored salivary flow rates to normal levels and significantly reduced the focus scores of SMGs in NOD mice. More than 1800 proteins in SMG cells were quantified by the proteomic approach. Many SMG proteins involved in inflammation and apoptosis were found to be down-regulated whereas those involved in salivary gland biology and development/regeneration were up-regulated in the BM Soup-treated mice. qPCR analysis also revealed expression changes of growth factors and cytokines in the SMGs of the treated NOD mice.ConclusionBM Soup treatment is effective to restore the function of damaged SGs in NOD mice. Through gene/protein expression analysis, we have found that BM Soup treatment might effectuate via inhibiting apoptosis, focal adhesion and inflammation whereas promoting development, regeneration and differentiation of the SG cells in NOD mice. These findings provide important insights on the potential mechanisms underlying the BM Soup treatment for functional restoration of damaged SGs in NOD mice. Additional studies are needed to further confirm the identified target genes and their related signaling pathways that are responsible for the BM Soup treatment.

Lyophilized bone marrow cell extract functionally restores irradiation‐injured salivary glands

“…The process to obtain this cell extract consists of three cycles of freeze-thawing at −80°C/+37°C to lyse the bone marrow cells. Then, the cell lysate is centrifuged at 17,000 g and the supernatant, named as BMCE or also BM Soup, is aliquoted and injected into the diseased animal (Fang et al, 2015;Misuno et al, 2014;Tran et al, 2013;Yeghiazarians et al, 2009). The native proteins were demonstrated to be the main effective ingredients in BMCE (Fang et al, 2015).…”

Identification of the active components in Bone Marrow Soup: a mitigator against irradiation-injury to salivary glands

“…These genes/pathways can be identified by using the gene network analysis of global expression alterations caused by BM Soup treatment. We have recently used a proteomics approach to quantify 1850 proteins, based on liquid chromatography with tandem mass spectrometry (LC-MS/MS) and tandem mass tagging (TMT), for global analysis of proteins in SG cells between BM Soup-treated and untreated NOD mice 22 . Many SG proteins in Sjögren’s-like disease mice were found to be altered by the BM Soup treatment.…”

“…Our group adapted this method and coined the term as “Bone Marrow Soup” ( BM Soup ), which represented all the yet-to-be-identified soluble components of the cell lysate from whole bone marrow cells. We reported that both whole BM cells and BM Soup restored comparable salivary function following IR and in Sjögren’s-like disease mice 21 22 . Because the BM Soup is an extract from a cell lysate, it is theoretically less immunogenic and tumorigenic 23 .…”

Potential Cell-Based Therapies for Irreversibly Damaged Salivary Glands and Atrophic Alveolar Bone

“…Thus, cell-based therapy must be recognized as one of the promising options for future clinical trials. Recently, our group has also shown that the injection of bone marrow cell extracts (a bioactive lysate of BMDCs) could restore the function of damaged salivary gland in NOD mice (Misuno et al, 2014).…”