A culture method with berbamine, a plant alkaloid, enhances CAR-T cell efficacy through modulating cellular metabolism.

Memory T cells demonstrate superior in vivo persistence and antitumor efficacy. However, methods for manufacturing less differentiated T cells are not yet well-established. Here, we show that producing chimeric antigen receptor (CAR)-T cells using berbamine (BBM), a natural compound found in the Chinese herbal medicine Berberis amurensis, enhances the antitumor efficacy of CAR-T cells. BBM is identified through cell-based screening of chemical compounds using induced pluripotent stem cell-derived T cells, leading to improved viability with a memory T cell phenotype. Transcriptomics and metabolomics using stem cell memory T cells reveal that BBM broadly enhances lipid metabolism. Furthermore, the addition of BBM downregulates the phosphorylation of p38 mitogen-activated protein kinase and enhanced mitochondrial respiration. CD19-CAR-T cells cultured with BBM also extend the survival of leukaemia mouse models due to their superior in vivo persistence. This technology offers a straightforward approach to enhancing the antitumor efficacy of CAR-T cells.

Mini-TCRs: Truncated T cell receptors to generate T cells from induced pluripotent stem cells.

Allogeneic T cell platforms utilizing induced pluripotent stem cell (iPSC) technology exhibit significant promise for the facilitation of adoptive immunotherapies. While mature T cell receptor (TCR) signaling plays a crucial role in generating T cells from iPSCs, the introduction of exogenous mature TCR genes carries a potential risk of causing graft-versus-host disease (GvHD). In this study, we present the development of truncated TCRα and TCRß chains, termed mini-TCRs, which lack variable domains responsible for recognizing human leukocyte antigen (HLA)-peptide complexes. We successfully induced cytotoxic T lymphocytes (CTLs) from iPSCs by employing mini-TCRs. Combinations of TCRα and TCRß fragments were screened from mini-TCR libraries based on the surface localization of CD3 proteins and their ability to transduce T cell signaling. Consequently, mini-TCR-expressing iPSCs underwent physiological T cell development, progressing from the CD4 and CD8 double-positive stage to the CD8 single-positive stage. The resulting iPSC-derived CTLs exhibited comparable cytokine production and cytotoxicity in comparison to that of full-length TCR-expressing T lymphocytes when chimeric antigen receptors (CARs) were expressed. These findings demonstrate the potential of mini-TCR-carrying iPSCs as a versatile platform for CAR T cell therapy, offering a promising avenue for advancing adoptive immunotherapies.

Effects of Vibration Therapy on Immobilization-Induced Hypersensitivity in Rats.

BACKGROUND: Cast immobilization induces mechanical hypersensitivity, which disturbs rehabilitation. Although vibration therapy can reduce various types of pain, whether vibration reduces immobilization-induced hypersensitivity remains unclear. OBJECTIVE: The purpose of this study was to investigate the preventive and therapeutic effects of vibration therapy on immobilization-induced hypersensitivity. DESIGN: The experimental design of the study involved conducting behavioral, histological, and immunohistochemical studies in model rats. METHODS: Thirty-five Wistar rats (8 weeks old, all male) were used. The right ankle joints of 30 rats were immobilized by plaster cast for 8 weeks, and 5 rats were used as controls. The immobilized rats were divided randomly into the following 3 groups: (1) immobilization-only group (Im, n=10); (2) vibration therapy group 1, for which vibration therapy was initiated immediately after the onset of immobilization (Im+Vib1, n=10); and (3) vibration therapy group 2, for which vibration therapy was initiated 4 weeks after the onset of immobilization (Im+Vib2, n=10). Vibration was applied to the hind paw. The mechanical hypersensitivity and epidermal thickness of the hind paw skin were measured. To investigate central sensitization, calcitonin gene-related peptide (CGRP) expression in the spinal cord and dorsal root ganglion (DRG) was analyzed. RESULTS: Immobilization-induced hypersensitivity was inhibited in the Im+Vib1 group but not in the Im+Vib2 group. Central sensitization, which was indicated by increases in CGRP expression in the spinal cord and the size of the area of CGRP-positive neurons in the DRG, was inhibited in only the Im+Vib1 group. Epidermal thickness was not affected by vibration stimulation. LIMITATIONS: A limitation of this study is that the results were limited to an animal model and cannot be generalized to humans. CONCLUSIONS: The data suggest that initiation of vibration therapy in the early phase of immobilization may inhibit the development of immobilization-induced hypersensitivity.

Hyperalgesia in an immobilized rat hindlimb: effect of treadmill exercise using non-immobilized limbs.

Cast immobilization of limbs causes hyperalgesia, which is a decline of the threshold of mechanical and thermal mechanical stimuli. The immobilization-induced hyperalgesia (IIH) can disturb rehabilitation and activities of daily living in patients with orthopedic disorders. However, it is unclear what therapeutic and preventive approaches can be used to alleviate IIH. Exercise that activates the descending pain modulatory system may be effective for IIH. The purpose of this study was to investigate the effects of treadmill exercise during the immobilization period, using the non-immobilized limbs, on IIH. Thirty-six 8-week-old Wistar rats were randomly divided into (1) control, (2) immobilization (Im), and (3) immobilization and treadmill exercise (Im+Ex) groups. In the Im and Im+Ex groups, the right ankle joints of each rat were immobilized in full plantar flexion with a plaster cast for an 8-week period. In the Im+Ex group, treadmill exercise (15 m/min, 30 min/day, 5 days/week) was administered during the immobilization period while the right hindlimb was kept immobilized. Mechanical hyperalgesia was measured using von Frey filaments every week. To investigate possible activation of the descending pain modulatory system, beta-endorphin expression levels in hypothalamus and midbrain periaqueductal gray were analyzed. Although IIH clearly occurred in the Im group, the hyperalgesia was partially but significantly reduced in the Im+Ex group. Beta-endorphin, which is one of the endogenous opioids, was selectively increased in the hypothalamus and midbrain periaqueductal gray of the Im+Ex group. Our data suggest that treadmill running using the non-immobilized limbs reduces the amount of hyperalgesia induced in the immobilized limb even if it is not freed. This ameliorating effect might be due to the descending pain modulatory system being activated by upregulation of beta-endorphin in the brain.

Immobilization-induced hypersensitivity associated with spinal cord sensitization during cast immobilization and after cast removal in rats.

This study examined mechanical and thermal hypersensitivity in the rat hind paw during cast immobilization of the hind limbs for 4 or 8 weeks and following cast removal. Blood flow, skin temperature, and volume of the rat hind paw were assessed in order to determine peripheral circulation of the hind limbs. Sensitization was analyzed by measuring the expression of the calcitonin gene-related peptide (CGRP) in the spinal dorsal horn following cast immobilization. Two weeks post immobilization, mechanical and thermal sensitivities increased significantly in all rats; however, peripheral circulation was not affected by immobilization. Cast immobilization for 8 weeks induced more serious hypersensitivity compared to cast immobilization for 4 weeks. Moreover, CGRP expression in the deeper lamina layer of the spinal dorsal horn increased in the rats immobilized for 8 weeks but not in those immobilized for 4 weeks. These findings suggest that immobilization-induced hypersensitivity develops during the immobilization period without affecting peripheral circulation. Our results also highlight the possibility that prolonged immobilization induces central sensitization in the spinal cord.