Quality of life-improving effect of autologous ex vivo expanded cytotoxic and opioid-producing lymphocytes for dogs with cancers.

Activated lymphocyte therapy is one of the immunotherapies for cancer patients that is expected to prolong life without any adverse effects and maintain satisfactory quality of life (QOL). However, the objective assessment and maintenance of a standardized evaluation of QOL are not easy. We aimed to evaluate activated autologous lymphocyte therapy for cancer dogs using the characteristics of the cultured cells and QOL as perceived by owners. In in vitro experiments, peripheral blood mononuclear cells (PBMCs) collected from healthy dogs were stimulated using anti-CD3 antibody and recombinant interleukin-2 under a closed system. The number of CD4+ and CD8+ T lymphocytes in the cultured cells was higher than that of PBMCs (P < 0.05). Natural killer activity, proenkephalin (known as the precursor of endogenous opioids) and interferon-γ mRNA in activated lymphocytes were significantly higher than in PBMCs (P < 0.05). Met-enkephalin was detected in activated lymphocytes. QOL of 58 dogs afflicted with common types of cancers in humans increased after every administration of activated lymphocyte therapy (P < 0.05). Overall, these results indicated that activated lymphocyte therapy could have beneficial effects on QOL in dogs with cancers. This was objectively evaluated and this improvement was related to presence of opioid-producing lymphocytes.

Long-Term Trypsin Treatment Promotes Stem Cell Potency of Canine Adipose-Derived Mesenchymal Stem Cells.

Mesenchymal stem cells (MSCs) isolated from adipose tissue (adipose-derived stem cells [ADSCs]) are considered one of the most promising cell types for applications in regenerative medicine. However, the regenerative potency of ADSCs may vary because of heterogeneity. Long-term trypsin treatment (LTT) is known to significantly concentrate multilineage-differentiating stress-enduring (Muse) cells from human MSCs. In this study, we aimed to generate cells with high stem cell potency from canine ADSCs using LTT. After 16 h of treatment with trypsin, surviving ADSCs (LTT-tolerant cells) had significantly enhanced expression of stage-specific embryonic antigen (SSEA)-1, a mouse embryonic stem cell marker, and fucosyltransferase 9, one of several fucosyltransferases for SSEA-1 biosynthesis. However, LTT-tolerant cells did not enhance the expression of SSEA-3, a known human Muse cell marker. LTT-tolerant cells, however, showed significantly higher self-renewal capacity in the colony-forming unit fibroblast assay than ADSCs. In addition, the LTT-tolerant cells formed cell clusters similar to embryoid bodies and expressed undifferentiated markers. Moreover, these cells differentiated into cells of all three germ layers and showed significantly higher levels of α 2-6 sialic acid (Sia)-specific lectins, known as differentiation potential markers of human MSCs, than ADSCs. LTT-tolerant cells had a normal karyotype and had low telomerase activity, showing little carcinogenetic potency. LTT-tolerant cells also showed significantly increased activity of transmigration in the presence of chemoattractants and had increased expression of migration-related genes compared with ADSCs. In addition, LTT-tolerant cells had stronger suppressive activity against mitogen-stimulated lymphocyte proliferation than ADSCs. Overall, these results indicated that the LTT-tolerant cells in canine ADSCs have similar properties as human Muse cells (although one of the undifferentiated markers is different) and are expected to be a promising tool for regenerative therapy in dogs.