An urgent need exists in clinical medicine for suitable alternatives to available techniques for bone tissue repair. Human adipose-derived stem cells (hASCs) represent a readily available, autogenous cell source with well-documented in vivo osteogenic potential. In this article, we manipulated Noggin expression levels in hASCs using lentiviral and nonintegrating minicircle short hairpin ribonucleic acid (shRNA) methodologies in vitro and in vivo to enhance hASC osteogenesis. Human ASCs with Noggin knockdown showed significantly increased bone morphogenetic protein (BMP) signaling and osteogenic differentiation both in vitro and in vivo, and when placed onto a BMP-releasing scaffold embedded with lentiviral Noggin shRNA particles, hASCs more rapidly healed mouse calvarial defects. This study therefore suggests that genetic targeting of hASCs combined with custom scaffold design can optimize hASCs for skeletal regenerative medicine.
CD8-expressing T cells are the main effector cells in cancer immunotherapy. Treatment-induced changes in intratumoral CD8 þ T cells may represent a biomarker to identify patients responding to cancer immunotherapy. Here, we have used a 89 Zr-radiolabeled human CD8-specific minibody (89 Zr-Df-IAB22M2C) to monitor CD8 þ T-cell tumor infiltrates by PET. The ability of this tracer to quantify CD8 þ T-cell tumor infiltrates was evaluated in preclinical studies following single-agent treatment with FOLR1-T-cell bispecific (TCB) antibody and combination therapy of CEA-TCB (RG7802) and CEA-targeted 4-1BB agonist CEA-4-1BBL. In vitro cytotoxicity assays with peripheral blood mononuclear cells and CEAexpressing MKN-45 gastric or FOLR1-expressing HeLa cervical cancer cells confirmed noninterference of the anti-CD8-PETtracer with the mode of action of CEA-TCB/CEA-4-1BBL and FOLR1-TCB at relevant doses. In vivo, the extent of tumor regression induced by combination treatment with CEA-TCB/ CEA-4-1BBL in MKN-45 tumor-bearing humanized mice correlated with intratumoral CD8 þ T-cell infiltration. This was detectable by 89 Zr-IAB22M2C-PET and g-counting. Similarly, single-agent treatment with FOLR1-TCB induced strong CD8 þ T-cell infiltration in HeLa tumors, where 89 Zr-Df-IAB22M2C again was able to detect CD8 tumor infiltrates. CD8-IHC confirmed the PET imaging results. Taken together, the anti-CD8minibody 89 Zr-Df-IAB22M2C revealed a high sensitivity for the detection of intratumoral CD8 þ T-cell infiltrates upon either single or combination treatment with TCB antibody-based fusion proteins. These results provide further evidence that the anti-CD8 tracer, which is currently in clinical phase II, is a promising monitoring tool for intratumoral CD8 þ T cells in patients treated with cancer immunotherapy. Significance: Monitoring the pharmacodynamic activity of cancer immunotherapy with novel molecular imaging tools such as 89 Zr-Df-IAB22M2C for PET imaging is of prime importance to identify patients responding early to cancer immunotherapy.
(1) Morphiceptin and PFRTic-NH played important roles in the regulation of gastrointestinal motility; (2) MCRT possessed higher bioactivity of pain relief than gastrointestinal regulation, suggesting its promising analgesic property; (3) MCRT-induced motility disorders were sensitive to DOR but not to MOR blockade, indicating the pain-relieving specificity of speculated MOR subtype or splice variant or MOR-DOR heterodimer.
Increasing evidence suggests that NODs are involved in liver diseases; however, the underlying mechanisms remain obscure. In the present study, we analyzed the effect of NOD1 agonist pretreatment on acute liver failure induced by lipopolysaccharide (LPS) in D-galactosamine (D-GalN)-sensitized mice. We found that pretreatment with the NOD1 agonist markedly reduced LPS/D-GalN-induced mortality, elevation of serum ALT levels, and hepatocyte apoptosis. The protective effect of NOD1 agonist was independent of tumor necrosis factor (TNF)-α inhibition. NOD1 agonist pretreatment also attenuated TNF-α/D-GalN-induced apoptotic liver damage. The anti-apoptotic protein A20 expression was more pronounced in NOD1 agonist pretreated mice than in controls, and knockdown of A20 abrogated the protective effect of NOD1 agonist on LPS/D-GalN-induced liver injury and hepatocyte apoptosis. Further experiments showed that NOD1 agonist-induced A20 upregulation required the presence of kupffer cells and TNF-α. Taken together, our data strongly indicate that NOD1 is involved in the regulation of liver injury and could be a potential therapeutic target for liver diseases.
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