Pharmacological inhibition of caspase and calpain proteases: a novel strategy to enhance the homing responses of cord blood HSPCs during expansion

• Published in: PloS one 2012-01, Vol.7 (1), p.e29383
• Main Authors: Sangeetha, V M, Kadekar, Darshana, Kale, Vaijayanti P, Limaye, Lalita S
• Format: Article
• Language: English
• Subjects: Actin; Actins - chemistry; Adhesion tests; Adhesives; Amino Acid Chloromethyl Ketones - pharmacology; Analysis; Animals; Apoptosis; Biology; Blood; Bone marrow; Bone marrow transplantation; Calpain; Calpain - antagonists & inhibitors; Caspase Inhibitors; CD34 antigen; Cell adhesion; Cell Adhesion - drug effects; Cell Adhesion Molecules - genetics; Cell Adhesion Molecules - metabolism; Cell cycle; Cell Movement - drug effects; Cell Polarity - drug effects; Cell Size - drug effects; Cells (biology); Chemokine CXCL12 - genetics; Chemokine CXCL12 - metabolism; Chemokines; College campuses; Cord blood; CXCR4 protein; Cysteine; Cytokines; Expansion; Female; Fetal Blood - cytology; Flow cytometry; Gene Expression Regulation - drug effects; Growth factors; Hematopoietic stem cells; Hematopoietic Stem Cells - cytology; Hematopoietic Stem Cells - drug effects; Hematopoietic Stem Cells - metabolism; Homing; Humans; Inhibition; Integrin alpha4 - metabolism; Integrin alpha5 - metabolism; Integrins; Kinases; Laboratories; Medicine; Mice; Muscle proteins; Pharmacology; Polymerization; Population; Priming; Progenitor cells; Protease inhibitors; Protease Inhibitors - pharmacology; Protein Multimerization - drug effects; Protein Structure, Quaternary; Proteinase inhibitors; Receptors, CXCR4 - genetics; Receptors, CXCR4 - metabolism; rho GTP-Binding Proteins - metabolism; Standard deviation; Stem cells; Transplantation; Transplants & implants
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0029383

Expansion of hematopoietic stem/progenitor cells (HSPCs) is a well-known strategy employed to facilitate the transplantation outcome. We have previously shown that the prevention of apoptosis by the inhibition of cysteine proteases, caspase and calpain played an important role in the expansion and engraftment of cord blood (CB) derived HSPCs. We hypothesize that these protease inhibitors might have maneuvered the adhesive and migratory properties of the cells rendering them to be retained in the bone marrow for sustained engraftment. The current study was aimed to investigate the mechanism of the homing responses of CB cells during expansion. CB derived CD34(+) cells were expanded using a combination of growth factors with and without Caspase inhibitor -zVADfmk or Calpain 1 inhibitor- zLLYfmk. The cells were analyzed for the expression of homing-related molecules. In vitro adhesive/migratory interactions and actin polymerization dynamics of HSPCs were assessed. In vivo homing assays were carried out in NOD/SCID mice to corroborate these observations. We observed that the presence of zVADfmk or zLLYfmk (inhibitors) caused the functional up regulation of CXCR4, integrins, and adhesion molecules, reflecting in a higher migration and adhesive interactions in vitro. The enhanced actin polymerization and the RhoGTPase protein expression complemented these observations. Furthermore, in vivo experiments showed a significantly enhanced homing to the bone marrow of NOD/SCID mice. Our present study reveals another novel aspect of the regulation of caspase and calpain proteases in the biology of HSPCs. The priming of the homing responses of the inhibitor-cultured HSPCs compared to the cytokine-graft suggests that the modulation of these proteases may help in overcoming the major homing defects prevalent in the expansion cultures thereby facilitating the manipulation of cells for transplant procedures.