The activation of oncogenic signaling pathways induces the reprogramming of glucose metabolism in tumor cells and increases lactic acid secretion into the tumor microenvironment. This is a well-known characteristic of tumor cells, termed the Warburg effect, and is a candidate target for antitumor therapy. Previous reports show that lactic acid secreted by tumor cells is a proinflammatory mediator that activates the IL-23/IL-17 pathway, thereby inducing inflammation, angiogenesis and tissue remodeling. Here, we show that lactic acid, or more specifically the acidification it causes, increases arginase I (ARG1) expression in macrophages to inhibit T-cell proliferation and activation. Accordingly, we hypothesized that counteraction of the immune effects by lactic acid might suppress tumor development. We show that dichloroacetate (DCA), an inhibitor of pyruvate dehydrogenase kinases, targets macrophages to suppress activation of the IL-23/IL-17 pathway and the expression of ARG1 by lactic acid. Furthermore, lactic acid-pretreated macrophages inhibited CD8 1 T-cell proliferation, but CD8 1 T-cell proliferation was restored when macrophages were pretreated with lactic acid and DCA. DCA treatment decreased ARG1 expression in tumor-infiltrating immune cells and increased the number of IFN-c-producing CD8 1 T cells and NK cells in tumor-bearing mouse spleen. Although DCA treatment alone did not suppress tumor growth, it increased antitumor immunotherapeutic activity of Poly(IC) in both CD8 1 T cell-and NK cell-sensitive tumor models. Therefore, DCA acts not only on tumor cells to suppress glycolysis but also on immune cells to improve the immune status modulated by lactic acid and to increase the effectiveness of antitumor immunotherapy.Many types of immune cells infiltrate tumors. Although these immune cells were classically thought to attack and eliminate tumors, recent studies indicate that they actually induce inflammation within tumors, thereby promoting tumor progression by inducing angiogenesis and tissue remodeling within the tumor microenvironment and tumor invasion and metastasis. 1,2 Furthermore, immune cells such as tumorassociated macrophages (TAMs), myeloid-derived suppressor cells (MDSCs) and regulatory T cells (Tregs), all of which have potent suppressive effects on anticancer immune responses, are also recruited to tumors. 3,4 We previously showed that lactic acid secreted by tumor cells enhances the production of IL-23 by monocytes/macrophages stimulated with Toll-like receptor (TLR) ligands. 5

Reprogramming of glucose metabolism in tumor cells is referred to as the Warburg effect and results in increased lactic acid secretion into the tumor microenvironment. We have previously shown that lactic acid has important roles as a pro‐inflammatory and immunosuppressive mediator and promotes tumor progression. In this study, we examined the relationship between the lactic acid concentration and expression of LDHA and GLUT1, which are related to the Warburg effect, in human head and neck squamous cell carcinoma (HNSCC). Tumors expressing lower levels of LDHA and GLUT1 had a higher concentration of lactic acid than those with higher LDHA and GLUT1 expression. Lactic acid also suppressed the expression of LDHA and GLUT1 in vitro. We previously reported that lactic acid enhances expression of an M2 macrophage marker, ARG1, in murine macrophages. Therefore, we investigated the relationship between the lactic acid concentration and polarization of M2 macrophages in HNSCC by measuring the expression of M2 macrophage markers, CSF1R and CD163, normalized using a pan‐macrophage marker, CD68. Tumors with lower levels of CD68 showed a higher concentration of lactic acid, whereas those with higher levels of CSF1R showed a significantly higher concentration of lactic acid. A similar tendency was observed for CD163. These results suggest that tumor‐secreted lactic acid is linked to the reduction of macrophages in tumors and promotes induction of M2‐like macrophage polarization in human HNSCC.

In the reflexively standing acute decerebrate cat, we have previously shown that pulse train microstimulation of the hook bundle of Russel in the midline of the cerebellar white matter, through which crossed fastigiofugal fibers decussate, augments the postural tone of neck, trunk, fore-, and hindlimb extensor muscles. In the present study we examined the possible role of such stimulation in evoking locomotion as the animal is supported by a rubber hammock with its feet contacting the moving surface of a treadmill. We were able to provoke well-coordinated, bilaterally symmetrical, fore- and hindlimb movements, whose cycle time and pattern were controlled by appropriate changes in stimulus intensity and treadmill speed. We carefully and systematically mapped this cerebellar locomotor region (CLR) through repeated dorsoventral penetrations with a glass-coated tungsten microelectrode in a single animal and between animals. We found that the optimal locus for evoking locomotion was centered on the midline, at Horsley-Clarke coordinates H0 and P7.0, and extended over a rostrocaudal and dorsolateral range of approximately 0.5 mm. The lowest effective stimulus intensity at the optimal site was in the range of 5-8 microA. Along penetration tracks to left or right of the midline, effective stimulus intensity increased and evoked locomotor patterns were no longer symmetrical, but rather shifted toward the contralateral limbs. In the same animals, controlled locomotion was evoked by stimulating the mesencephalic locomotor region (MLR). With concomitant stimulation of the optimal sites in the CLR and the MLR, each at subthreshold strength, locomotor movements identical to those seen with suprathreshold stimulation of each site alone were evoked. With concomitant stimulation at suprathreshold strength for each site, locomotion became vigorous, with a shortened cycle time. After making ablative lesions at either the CLR or MLR (unilateral or bilateral), controlled locomotion was still evoked at the prior stimulus strength by stimulating the remaining site. Together, these results demonstrate that selective stimulation of the hook bundle of Russel in the midsagittal plane of the cerebellar white matter evokes "controlled" locomotion identical to that evoked by stimulating the MLR. We have shown that the fastigial nucleus is one of the supraspinal locomotion inducing sites and that it can independently and simultaneously trigger brain stem and spinal locomotor subprograms formerly believed to be the domain of various brain stem regions including the MLR and the subthalamic locomotor region.

In a decerebrate cat (locomotor preparation), stimulation of a restricted region along the midline cerebellar white matter has been found to evoke generalized augmentation of postural muscle tone on a stationary surface (Asanome et al. 1998. Neurosci. Res. 30: 257-269) and "controlled" locomotion on the surface of a moving treadmill. Characteristics of cerebellar-evoked locomotion were similar to those of mesencephalic locomotor region-evoked "controlled" locomotion on the same animal. Microinjection of a neural tracer (CTb-HRP) into the lesioned stimulus site of the cerebellar white matter resulted in both retrograde labelling of cells in the fastigial nuclei, bilaterally, and anterograde labeling of fibers descending to the brain stem. These results indicated that the effective cerebellar stimulus site (cerebellar locomotor region) corresponded to the midline region of the hook bundle of Russell (Rasmussen, A. T., 1933. J. Comp. Neurol. 57: 165-197), through which crossed fastigioreticular, fastigiovestibular, and fastigiospinal fibers pass. In this study, contribution of reticulospinal systems to the control of cerebellar-evoked locomotion was extensively studied. By stimulating the cerebellar locomotor region and the MLR in the same animal, a majority of antidromically identified pontomedullary reticulospinal cells were synaptically activated. The results of the present study demonstrated that fastigial cells with crossed fastigioreticular fibers and reticulospinal fibers play a crucial role in the control of posture and locomotion in the locomotor preparation.

The faithful shutdown of the somatic program occurs in the early stage of reprogramming. Here, we examined the effect of in vivo reprogramming on Kras-induced cancer development. We show that the transient expression of reprogramming factors (1–3 days) in pancreatic acinar cells results in the transient repression of acinar cell enhancers, which are similarly observed in pancreatitis. We next demonstrate that Kras and p53 mutations are insufficient to induce ERK signaling in the pancreas. Notably, the transient expression of reprogramming factors in Kras mutant mice is sufficient to induce the robust and persistent activation of ERK signaling in acinar cells and rapid formation of pancreatic ductal adenocarcinoma. In contrast, the forced expression of acinar cell-related transcription factors inhibits the pancreatitis-induced activation of ERK signaling and development of precancerous lesions in Kras-mutated acinar cells. These results underscore a crucial role of dedifferentiation-associated epigenetic regulations in the initiation of pancreatic cancers.