Optimized CAR-T therapy based on spatiotemporal changes and chemotactic mechanisms of MDSCs induced by hypofractionated radiotherapy.

Poor intratumoral infiltration is the major challenge for chimeric antigen receptor (CAR)-T cell therapy in solid tumors. Hypofractionated radiotherapy (HFRT) has been reported to induce immune cell infiltration and reshape the tumor immune microenvironment. Here, we showed that HFRT (5 × 5 Gy) mediated an early accumulation of intratumoral myeloid-derived suppressor cells (MDSCs) and decreased infiltration of T cells in the tumor microenvironment (TME) of immunocompetent mice bearing triple-negative breast cancer (TNBC) or colon cancer, which was further confirmed in tumors from patients. RNA sequencing (RNA-seq) and cytokine profiling analysis revealed that HFRT induced the activation and proliferation of tumor-infiltrated MDSCs, which was mediated by the interactions of multiple chemokines and chemokine receptors. Further investigation showed that when combined with HFRT, CXCR2 blockade significantly inhibited MDSCs trafficking to tumors and effectively enhanced the intratumoral infiltration and treatment efficacy of CAR-T cells. Our study demonstrates that MDSCs blockade combined with HFRT is promising for CAR-T cell therapy optimization in solid tumors.

A Secure and Efficient Data Sharing and Searching Scheme in Wireless Sensor Networks.

Wireless sensor networks (WSN) generally utilize cloud computing to store and process sensing data in real time, namely, cloud-assisted WSN. However, the cloud-assisted WSN faces new security challenges, particularly outsourced data confidentiality. Data Encryption is a fundamental approach but it limits target data retrieval in massive encrypted data. Public key encryption with keyword search (PEKS) enables a data receiver to retrieve encrypted data containing some specific keyword in cloud-assisted WSN. However, the traditional PEKS schemes suffer from an inherent problem, namely, the keyword guessing attack (KGA). KGA includes off-line KGA and on-line KGA. To date, the existing literature on PEKS cannot simultaneously resist both off-line KGA and on-line KGA performed by an external adversary and an internal adversary. In this work, we propose a secure and efficient data sharing and searching scheme to address the aforementioned problem such that our scheme is secure against both off-line KGA and on-line KGA performed by external and internal adversaries. We would like to stress that our scheme simultaneously achieves document encryption/decryption and keyword search functions. We also prove our scheme achieves keyword security and document security. Furthermore, our scheme is more efficient than previous schemes by eliminating the pairing computation.

[The Construction of ROR1 Targeting Chimeric Antigen Receptor Modified T Cells and Its Killing Effect for ROR1-positive Tumor Cells].

OBJECTIVE: To test the killing effect of type Ⅰ receptor tyrosine kinase-like orphan receptor (ROR1) chimeric antigen receptor T cell (CAR-T) on several ROR1-expressing tumor cells in vitro. METHODS: The CAR gene was designed and synthesized by constructing the lentiviral vector plasmid, and BamHⅠ/EcoRⅠ was used to identify the plasmid. The expression levels of ROR1 among a variety of tumor cell lines were compared using flow cytometry (FCM). The killing effect of CAR-T on positive cells was detected by FCM, the LDH assay and ELISA. RESULTS: The double enzyme digestion identified CAR gene was successfully constructed to the lentivirus vector plasmid. FCM detection showed that the efficiency of CAR-T infection was about 47.23%. Multiple tumor cells expressed ROR1 in varying degrees. The FCM and the LDH assay indicated that CAR-T specifically killed ROR1-positive tumor cells. On positive target cells, more interferonI-γ (FN-γ) could be released during the CAR-T killing process than control T (P<0.05). CONCLUSION: We successfully constructed ROR1 CAR-T. CAR-T can specifically kill ROR1-positive tumor cells and cause the release of large amounts of IFN-γ, providing an experimental basis for clinical application.

PSME3 induces epithelial-mesenchymal transition with inducing the expression of CSC markers and immunosuppression in breast cancer.

Proteasome activator subunit 3 (PSME3) plays a key role in breast cancer by regulating the cell cycle. However, its role in other pathogenesis-related features of breast cancer is unclear. In this study, we found that overexpression of PSME3 induced the epithelial-mesenchymal transition and contributed to induce the expression of cancer stem cell markers of the MDA-MB-231 cell line, thus increasing the migration, and invasion of the cells. Moreover, overexpression of PSME3 reduced the chemotaxis of CD8+ T cells and induced the apoptosis of T cells in vitro. Furthermore, PSME3 knockdown increased the number of CD8+ T cells in vivo and reduced the subcutaneous tumor growth rate. These findings revealed that PSME3 induces epithelial-mesenchymal transition with inducing the expression of CSC markers and influencing the tumor immune microenvironment in breast cancer.

Malignant Pleural Effusion and ascites Induce Epithelial-Mesenchymal Transition and Cancer Stem-like Cell Properties via the Vascular Endothelial Growth Factor (VEGF)/Phosphatidylinositol 3-Kinase (PI3K)/Akt/Mechanistic Target of Rapamycin (mTOR) Pathway.

Malignant pleural effusion (PE) and ascites, common clinical manifestations in advanced cancer patients, are associated with a poor prognosis. However, the biological characteristics of malignant PE and ascites are not clarified. Here we report that malignant PE and ascites can induce a frequent epithelial-mesenchymal transition program and endow tumor cells with stem cell properties with high efficiency, which promotes tumor growth, chemoresistance, and immune evasion. We determine that this epithelial-mesenchymal transition process is mainly dependent on VEGF, one initiator of the PI3K/Akt/mechanistic target of rapamycin (mTOR) pathway. From the clinical observation, we define a therapeutic option with VEGF antibody for malignant PE and ascites. Taken together, our findings clarify a novel biological characteristic of malignant PE and ascites in cancer progression and provide a promising and available strategy for cancer patients with recurrent/refractory malignant PE and ascites.

Extravascular red blood cells and hemoglobin promote tumor growth and therapeutic resistance as endogenous danger signals.

Hemorrhage is a common clinical manifestation in patients with cancer. Intratumor hemorrhage has been demonstrated to be a poor prognostic factor for cancer patients. In this study, we investigated the role of RBCs and hemoglobin (Hb) in the process of tumor progression and therapeutical response. RBCs and Hb potently promoted tumor cell proliferation and syngenic tumor growth. RBCs and Hb activated the reactive oxygen species-NF-κB pathway in both tumor cells and macrophages. RBCs and Hb also induced chemoresistance mediated, in part, by upregulating ABCB1 gene expression. Tumor growth induced by RBCs was accompanied by an inflammatory signature, increased tumor vasculature, and influx of M2 macrophages. In both the peritoneal cavity and tumor microenvironment, extravascular RBCs rapidly recruited monocyte-macrophages into the lesion sites. In addition, RBCs and Hb increased several nucleotide-binding oligomerization domain-like receptors' expression and induced IL-1ß release. Our results provide novel insights into the protumor function of RBCs and Hb as endogenous danger signals, which can promote tumor cell proliferation, macrophage recruitment, and polarization. Hemorrhage may represent a useful prognostic factor for cancer patients because of its role in tumor promotion and chemoresistance.

EML4-ALK induces epithelial-mesenchymal transition consistent with cancer stem cell properties in H1299 non-small cell lung cancer cells.

The echinoderm microtubule-associated protein-like 4(EML4)--anaplastic lymphoma kinase (ALK) fusion gene has been identified as a driver mutation in non-small-cell lung cancer (NSCLC). However, the role of EML4-ALK in malignant transformation is not entirely clear. Here, for the first time, we showed that H1299 NSCLC cells stably expressing EML4-ALK acquire EMT phenotype, associated with enhanced invasive migration and increased expression of EMT-inducing transcription factors. H1299-EML4-ALK cells also displayed cancer stem cell-like properties with a concomitant up-regulation of CD133 and enhanced ability of mammospheres formation. Moreover, we found that inhibition of ERK1/2 reversed EMT induced by EML4-ALK in H1299 cells. Taken together, these results suggested that EML4-ALK induced ERK activation is mechanistically associated with EMT phenotype. Thus, inhibition of ERK signaling pathway could be a potential strategy in treatment of NSCLC patients with EML4-ALK translocation.

[REGgamma promotes malignant behaviors of lung cancer cells].

OBJECTIVE: To determine the expression of proteasome aotivator gamma (REGgamma) in human lung cancer tissues and cell lines and its association with malignant biological behaviors. METHODS: Immunohistochemistry (IHC) was used to detect the expression of REGgamma in lung cancer and normal lung tissues. The expressions of REGgamma in lung cancer cells and normal epithelial cells were determined by Western blot. The H1975 lung cancer stable cell lines with different levels of REGgamma expression were constructed and their proliferations were evaluated by MTT assay. PI staining was used to assess the influence of REGgamma on cell growth cycle. The effect of REGgamma on the migration of lung cancer cells were observed with the cell scratch experiment. RESULTS: Lung cancer tissues had significantly higher levels of REGgamma expression than normal tissues. Similarly, lung cancer cell lines showed higher levels of REGgamma expression than the normal epithelial cell line. The overexpression of REGgamma enhanced cancer cell proliferations (P < 0.05), promoted more cells into the S+G2/M phase (P < 0.05) and promoted the migration of cancer cells (P < 0.05). All of these effects were reversed after suppression of REGgamma. CONCLUSION: REGgamma facilitates malignant biological behaviors of lung cancers.

Enhanced interaction between natural killer cells and lung cancer cells: involvement in gefitinib-mediated immunoregulation.

BACKGROUND: Natural killer (NK) cells can kill tumor cells in a non-MHC-restricted manner. However, cancer cells frequently escape from the attack of NK cells by multiple ways. In this study, we investigated the effect of gefitinib on the interaction between NK cells and lung cancer cells. METHODS: 5¹Cr release assay, CD107a assay, and IFN-γ secretion assay were performed to detect the sensitivity of lung cancer cell lines A549 and H1975 to NK cells cytotoxicity in the presence of gefitinib. Human NK cells were co-cultured with A549 and H1975 cell lines in the presence of gefitinib. NKG2D ligands, ULBP1, ULBP2, MICA, and MHC-I on tumor cells, and NKG2D, NKp44 and NKp46 on NK cells were evaluated with flow cytometry. 51Cr release assay was performed when NKG2D antibody were added into the co-culture system. Expressions of stat3 and LC3 I/II on tumor cells were determined with western blot after co-cultured with NK cells. After treated with gefitinib, mannose-6-phosphate receptor (MPR) on H1975 cells was evaluated by flow cytometry. 5¹Cr release assay were performed when MPR antagonist were used. RESULTS: Gefitinib increased cytotoxicity of NK cells to human lung cancer H1975 cells with EGFR L858R + T790M mutations, while not in A549 cells with wild type EGFR. Gefitinib could block the immune escape by up-regulating the expression of NKG2D ligands ULBP1, ULBP2 or MICA on tumor cells and NKG2D on NK cells in the co-culture system. Gefitinib and NK cells up-regulated MHC-I expression in A549 while not in H1975 cells. NKG2D antibody blocked the enhanced NK cytotoxicity by gefitinib. The combination of NK cells and gefitinib could significantly down-regulate stat3 expression. Furthermore, NK cells-mediated tumor cell autophagy was observed in A549 cells while not in H1975 cells. Notably, gefitinib increased autophagy and MPR expression in H1975 cells, which improved the sensitivity to NK cell-based immunotherapy. CONCLUSIONS: Gefitinib greatly enhanced NK cell cytotoxicity to lung cancer cells with EGFR L858R + T790M resistance mutation. Combination of EGFR tyrokinase inhibitors and NK cells adoptive immunotherapy may represent a potentially effective strategy for patients with non-small cell lung cancer.

[Study on the anti-Lewis lung carcinoma effect of metformin combined with MCT1 inhibitor CHC].

OBJECTIVE: To investigate the antitumor effect of the combination of metformin with alpha-cyano-4-hydroxycinnamic acid (CHC, a MCT1 inhibitor) in the treatment of Lewis lung cancer. METHODS: In vitro, the utilization of lactate acid was measured by lactate assay in cultured medium and the inhibition of LL/2 cell proliferation of four groups [control group, metformin group (1 mmol/L and 5 mmol/L), CHC group (5 mmol/L) and the combination group (metformin 5 mmol/L and CHC 5 mmol/L)] was detected in 24 h, 48 h, and their apoptosis in 24 h was also detected. In vivo, twenty eight C57BL/6 mice bearing LL/2 (5 x 10(5)) subcutaneous Lewis lung cancer on the right flank was established and then randomly assigned into four groups: control, metformin (200 mg/kg body mass in 0.1 mL i. g. with NS 0.1 mL i. p.), CHC (100 mg/kg body mass in 0.1 mL i.p. with NS 0.1 mL i. g. ) and the combination (metformin 200 mg/kg body mass in 0.1 mL i.g. with CHC 100 mg/kg body mass in 0.1 mL i. p.). Tumor volume was measured. The pathologic observation and apoptotic analysis of tumors was assessed by TUNEL assay. RESULTS: Compared to the contorl, metformin or CHC alone, combination of two drugs leaded to a significant lactate acid production in cultured medium and the inhibition of LL/ 2 cell viability (P < 0.05). In vivo, the systemic administration of two drugs leaded to obvious retarded tumor growth compared with used alone in early stage (P < 0.05). The TUNEL assay showed the significantly increased number of apoptosis cells in tumor tissues from the combination group. CONCLUSION: Combination of metformin and CHC transformed the lactic acid metabolism in LL/2 cells and induced cell apoptosis and showed the antitumor effect.

A PD-L1-targeting chimeric switch receptor enhances efficacy of CAR-T cell for pleural and peritoneal metastasis.

Pleural and peritoneal metastasis accompanied by malignant pleural effusion (MPE) or malignant ascites (MA) is frequent in patients with advanced solid tumors that originate from the lung, breast, gastrointestinal tract and ovary. Regional delivery of CAR-T cells represents a new strategy to control tumor dissemination in serous cavities. However, malignant effusions constitute an immune-suppressive environment that potentially induces CAR-T cell dysfunction. Here, we demonstrated that the anti-tumor cytotoxicity of conventional 2nd-generation CAR-T cells was significantly inhibited by both the cellular and non-cellular components of MPE/MA, which was primarily attributed to impaired CAR-T cell proliferation and cytokine production in MPE/MA environment. Interestingly, we found that PD-L1 was widely expressed on freshly-isolated MPE/MA cells. Based on this feature, a novel PD-L1-targeting chimeric switch receptor (PD-L1.BB CSR) was designed, which can bind to PD-L1, switching the inhibitory signal into an additional 4-1BB signal. When co-expressed with a 2nd-generation CAR, PD-L1.BB CSR-modified CAR-T cells displayed superior fitness and enhanced functions in both culture medium and MPE/MA environment, causing rapid and durable eradication of pleural and peritoneal metastatic tumors in xenograft models. Further investigations revealed elevated expressions of T-cell activation, proliferation, and cytotoxicity-related genes, and we confirmed that PD-L1 scFv and 4-1BB intracellular domain, the two important components of PD-L1.BB CSR, were both necessary for the functional improvements of CAR-T cells. Overall, our study shed light on the clinical application of PD-L1.BB CSR-modified dual-targeting CAR-T cells. Based on this study, a phase I clinical trial was initiated in patients with pleural or peritoneal metastasis (NCT04684459).

Preclinical Safety Evaluation of a Recombinant Plasmid Vector Encoding Mature Human Neutrophil Peptide-1 by Repeated Local Administrations in Nonhuman Primates.

In our previous studies, a novel gene therapy approach was developed based on a plasmid vector pSecTag2B in which recombinant HNP1 gene was regulated under a cytomegalovirus promoter to encode a mature human neutrophil peptide-1 (HNP1) form. We showed for the first time in various tumor models, including human cancer xenografts, that overexpression of HNP1 in the tumor milieu by intratumoral pSecTag-HNP1 (pHNP1) administration efficiently attenuated in vivo tumor progression, mediated host immune responses to tumors, and produced a synergistic effect when combined with chemotherapeutics. In this study, a preclinical safety investigation of HNP1 gene therapy was conducted in nonhuman primates. Eleven cynomolgus monkeys were divided into three groups of three to four animals each and received either repeated s.c. injections of pHNP1/cationic liposome complexes at a low (0.625 mg/kg) or a high (2.5 mg/kg) dose or glucose as control. Significant HNP1 in vivo accumulation was detected after consecutive administrations. All primates reached the end of the study with good body conditions. Injection site inflammation was the only obvious toxic reaction during observation period. In addition, elevation of monocyte/macrophage and neutrophil as well as decline of lymphocyte were detected in the peripheral blood of pHNP1-treated primates. These alterations were partially alleviated at the end of observation period. Besides, dose-related histopathological changes of the immune organs were observed at necropsy, including a minimal thymic lymphocyte decrease and a minimal-to-mild lymph node erythrocyte increase, but which cannot be excluded from HNP1-induced immune reactions. Together, these data support future clinical studies of pHNP1-based local gene delivery in tumor patients.

CXCL13 expression in mouse 4T1 breast cancer microenvironment elicits antitumor immune response by regulating immune cell infiltration.

Breast cancer is the most commonly diagnosed cancer type and the leading cause of cancer-related deaths among women worldwide. Previous studies have reported contradictory performance of chemokine CXC motif ligand 13 (CXCL13) in breast cancer. In this study, The Cancer Genome Atlas database analysis revealed that CXCL13 was overexpressed in various human cancers including breast carcinoma, and associated with good clinical prognosis in breast cancer. Flow cytometry detection also found upregulated intracellular CXCL13 expression in human breast cancer cell lines. To explore the possible role of CXCL13 in the breast cancer microenvironment, mouse triple negative breast cancer (TNBC) was lentivirally transfected to stably overexpress mouse CXCL13 (4T1-CXCL13). Both parental 4T1 and 4T1-CXCL13 strains showed no in vitro or in vivo endogenous cell surface CXCR5 expression. In immune-competent BALB/c mice, the in vivo tumor growth of 4T1-CXCL13 was significantly inhibited and even completely eradicated, accompanied with increased infiltrations of CD4+, CD8+ T lymphocytes and CD11b+CD11c+ DCs. Further investigations showed that CXCL13 expression in the 4T1 tumor microenvironment elicited long-term antitumor immune memory, and rejection of distal parental tumor. The antitumor activity of CXCL13 was remarkedly impaired in BALB/cA-nu nude mice, or in BALB/c mice with CD8+ T lymphocyte or NK cell depletion. Our investigation indicated that CXCL13 expression in TNBC triggered effective antitumor immunity by chemoattracting immune cell infiltrations and could be considered as a novel prognostic marker for TNBC.

Silencing of pkm2 increases the efficacy of docetaxel in human lung cancer xenografts in mice.

Tumor aerobic glycolysis, or the Warburg effect, plays important roles in tumor survival, growth, and metastasis. Pyruvate kinase isoenzyme M2 (PKM2) is a key enzyme that regulates aerobic glycolysis in tumor cells. Recent research has shown that PKM2 can be used as a tumor marker for diagnosis and, in particular, as a potential target for cancer therapy. We investigated the effects of combining shRNA targeting PKM2 and docetaxel on human A549 lung carcinoma cells both in vivo and in vitro. We observed that the shRNA can significantly downregulate the expression level of PKM2. The decrease of PKM2 resulted in a decrease in ATP synthesis, which caused intracellular accumulation of docetaxel. Furthermore, the combination of pshRNA-pkm2 and docetaxel inhibited tumor growth and promoted more cancer cell apoptosis both in vivo and in vitro. Our findings suggest that targeting tumor glycolysis can increase the efficacy of chemotherapy. In particular, the targeting of PKM2 could, to some extent, be a new way of reversing chemotherapy resistance to cancer therapy.

Low-Dose Radiation Therapy Promotes Radiation Pneumonitis by Activating NLRP3 Inflammasome.

PURPOSE: To determine the role of NLRP3 inflammasome activation in low-dose radiation-induced radiation pneumonitis (RP) and to assess whether inhibition or deletion of the NLRP3 inflammasome is critical for conferring protection against RP. METHODS AND MATERIALS: The human monocytic THP-1 cells were treated with increasing doses of radiation to assess the activation of NLRP3 by Western blot and enzyme-linked immunosorbent assay. Reactive oxygen species (ROS) production was measured by flow cytometry, with or without ROS inhibitor treatment. A mouse thoracic radiation model that received different doses of radiation was used, and the lung tissues of thoracic-irradiated nlrp3-/- and wild-type C57BL/6 mice were examined by hematoxylin and eosin and immunofluorescence staining. The concentrations of cytokines in the bronchoalveolar lavage fluid were measured by enzyme-linked immunosorbent assay and Luminex multiplex assays. Lipopolysaccharide (LPS) was administered intranasally 28 days after thoracic irradiation, and NLRP3 inhibitor, MCC950 was administered intraperitoneally after irradiation at 2 different doses. RESULTS: (1) The NLRP3 inflammasome was activated in 2 Gy irradiated THP-1 cells; NLRP3 and cleaved-caspase-1 levels were not associated with dose escalation of irradiation. (2) Activation of the NLRP3 inflammasome was mediated by ROS, and ROS inhibitor treatment decreased the production of IL-1ß and IL-18 in vitro. (3) NLRP3 was activated in mouse lungs by irradiation at 2 Gy, 4 Gy, and 16 Gy, and NLRP3 activation was continuous for 8 weeks. (4) NLRP3 deletion protects against LPS-mediated monocyte infiltration in the mouse lung. (5) The administration of MCC950 decreased the inflammation score of the mice irradiated with 2 Gy or 16 Gy in vivo. CONCLUSIONS: Our results indicate that the NLRP3 inflammasome is activated by low-dose irradiation both in vitro and in vivo. Inhibition or deletion of NLRP3 can specifically alleviate the mouse lung inflammation caused by radiation and LPS treatment. This study reveals the mechanism of low-dose radiation therapy-induced RP and offers a possible treatment strategy.

Author Correction: A whole-cell tumor vaccine modified to express fibroblast activation protein induces antitumor immunity against both tumor cells and cancer-associated fibroblasts.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Continuous targeted kinase inhibitors treatment induces upregulation of PD-L1 in resistant NSCLC.

First-generation epidermal growth factor receptor (EGFR) targeted kinase inhibitors (TKIs) are still used in selected non-small cell lung cancer (NSCLC) patients despite the resistance. Based on the correlation of programmed cell death receptor ligand 1 (PD-L1) and EGFR signaling pathway, whether continuous TKIs treatment will affect PD-L1 expression after disease progression remains unclear. To investigate the potential change of PD-L1 expression in TKI-resistant NSCLC after continuous TKIs treatment, we treated H1975 and HCC827 for more than one month and explored the possible effect on immune cells as well as underlying biological mechanisms. We found that continuous exposure to TKIs induced upregulation of PD-L1 in H1975 and HCC827. Moreover, PD-L1 upregulation significantly inhibited proliferation and slightly promoted apoptosis of T cells. We observed the activation of STAT3 and ERK1/2 along with the PD-L1 upregulation. With the pathway inhibitors, we found ERK1/2 pathway involved in inducing PD-L1 in resistant lung cancer. This study provides preclinical evidence that continuous TKIs treatment may induce PD-L1 expression in resistant NSCLC, resulting in the suppression of T cell function and immune escape. ERK1/2 pathway inhibitors, PD-L1/PD-1 inhibitors or combination strategies should be considered to reverse the resistance to TKIs in NSCLC patients.

Vascular endothelial growth receptor 1 acts as a stress-associated protein in the therapeutic response to thalidomide.

Thalidomide (THD) exhibits antitumor effects in several types of cancer. However, the failure of THD to inhibit tumor growth has also been observed in a number of murine models in vivo. The mechanism involved in the therapeutic failure of THD remains unclear. The present study demonstrated that, accompanied by growth-arresting and apoptosis-inducing effects (P<0.05), THD upregulated vascular endothelial growth factor receptor 1 (VEGFR1) expression levels in CT26 murine colorectal carcinoma cell lines. This in vitro phenomenon was also observed in various other cell lines, including human umbilical vein endothelial cells, SW480, SW620 and HCT116. Reactive oxygen species (ROS) levels were increased compared with those in the untreated control when cells were exposed to THD (P<0.05). Furthermore, results suggested that ROS suppression may have provoked the induction of VEGFR1 expression to some extent. In addition, the results revealed that THD failed to inhibit CT26 tumor growth in vivo and the expression of VEGFR1 protein was elevated by THD treatment compared with the control group in the murine colorectal tumor model (P<0.05). The results of further experiments suggested that VEGFR1 was elevated in response to various stress-associated situations, including chemotherapy, radiotherapy and thermotherapy, which indicate that it may act as a stress-associated protein. The present findings provide a foundation for the future study of VEGFR1-targeted therapy to enhance the efficacy of current therapies.

The Promotion of Erythropoiesis via the Regulation of Reactive Oxygen Species by Lactic Acid.

The simultaneous increases in blood lactic acid and erythrocytes after intense exercise could suggest a link between lactate and the erythropoiesis. However, the effects of lactic acid on erythropoiesis remain to be elucidated. Here, we utilized a mouse model to determine the role of lactic acid in this process in parallel with studies using leukaemic K562 cells. Treatment of K562 cells in vitro with lactic acid increased the mRNA and protein expression of haemoglobin genes and the frequency of GPA+ cells. Also, increases in haematocrit and CD71-/Ter119+ erythroid cells were observed in lactic acid-treated mice, which showed a physiological increase in blood lactate. Mouse bone marrow CD34+/CD117- cells showed an increase in erythroid burst-forming units after stimulation with lactic acid in vitro. Furthermore, lactic acid increased the intracellular reactive oxygen species (ROS) content in bone marrow and in K562 cells. Erythroid differentiation induced in Haematopoietic Stem Cells (HSCs) and K562 cells by lactic acid was abolished by reducing ROS levels with SOD or 2-mercaptoethanol, which suggests that ROS is a critical regulator of this process. These findings provide a better understanding of the role of lactic acid in cellular metabolism and physiological functions.

Corrigendum: A whole-cell tumor vaccine modified to express fibroblast activation protein induces antitumor immunity against both tumor cells and cancer-associated fibroblasts.

This corrects the article DOI: 10.1038/srep14421.