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ARS Home » Southeast Area » Little Rock, Arkansas » Arkansas Children's Nutrition Center » Microbiome and Metabolism Research » Research » Publications at this Location » Publication #421778
Research Project: Early Life Factors and Microbiota Impact on Healthy Development

Location: Microbiome and Metabolism Research

Title: Bone tissue specific delivery of Cystatin E/M proteins using Clostridial collagenase collagen binding domain

Author
item CAVINESS, PERRY - Arkansas Children'S Nutrition Research Center (ACNC)
item LAZARENKO, OXANA - Arkansas Children'S Nutrition Research Center (ACNC)
item XU, HONGWEI - University Arkansas For Medical Sciences (UAMS)
item BLACKBURN, MICHAEL - Arkansas Children'S Nutrition Research Center (ACNC)
item ZHAN, FENGHUANG - University Arkansas For Medical Sciences (UAMS)
item CHEN, JIN-RAN - Arkansas Children'S Nutrition Research Center (ACNC)

Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: 7/2/2024
Publication Date: N/A
Citation: N/A

Interpretive Summary:

Technical Abstract: Previously our group found that Cystatin E/M (CST6) ameliorates increased bone resorption in both multiple myeloma (MM) and sex steroid deficient (OVX) mouse models comparable to the bisphosphonate zoledronic acid and that increased estrogen influx may be responsible to CST6’s bone protective ability. mRNA microarray of MM tumor cells coupled with PER-CT analysis of newly diagnosed MM patients revealed that tumor cell expression of CST6 is associated with a decreased number of osteolytic lesions. Since CST6 is secreted by MM tumors, high concentrations may have harmful side effects. To avoid potential harmful side effects of CST6 on non-skeletal tissue we have generated a bone tissue targeting collagen binding domain (CBD)-CST6 fusion protein as well as a CBD-CST6 peptide (Asp71-Glu94). pET vectors were designed to fuse C-terminal ends of CST6 and CST6 peptide to N-terminal end of Clostridial collagenase H CBD and purchased commercially from VectorBuilder. Due to solubility concerns, CST6 peptide-CBD was expressed still containing N-terminal GST tag. CST6-CBD and CST6 peptide-CBD were found to be comparable to CST6 at suppressing osteoclastogenesis of both Raw 264.7 and mouse bone marrow primary cells. CST6-CBD, CST6 peptide-CBD and PBS (as control) were i.p. injected (200 µg/kg) into female C57BL/6 mice. Mice were sacrificed 5 hr. post injection, tissue (tibia and liver) was collected and slides were prepared for immunohistology. CST6-CBD and CST6 peptide-CBD injected were shown to have significantly increased amounts of CST6 present in only tibia and not in the other tissues collected. We have previously determined that CST6 treatment increases ERa protein concentration in tibia from OVX mice as well as ERa mRNA and protein levels in Raw 264.7 cells. Real-time PCR results for tibia from CST6-CBD or CST6 peptide-CBD injected revealed a significant increase in ERa mRNA levels. In addition, mRNA levels for TGFß as well as osteoclastogenesis promoters NFATc1 and NF'B in CST6-CBD injected mice were increased potentially suggesting CST6 suppress osteoclastogenesis through inhibition of SMAD signaling pathways. However, further research would be required to investigate this potential mechanism. Supported by in part by funding from USDA-ARS Project 6026-51000-010-05S; and NIH R01 project R37 AA18282 sub-awarded to JRC.