Caspase 8 deletion causes infection/inflammation-induced bone marrow failure and MDS-like disease in mice.

Myelodysplastic syndromes (MDS) are a heterogeneous group of pre-leukemic hematopoietic disorders characterized by cytopenia in peripheral blood due to ineffective hematopoiesis and normo- or hypercellularity and morphologic dysplasia in bone marrow (BM). An inflammatory BM microenvironment and programmed cell death of hematopoietic stem/progenitor cells (HSPCs) are thought to be the major causes of ineffective hematopoiesis in MDS. Pyroptosis, apoptosis and necroptosis (collectively, PANoptosis) are observed in BM tissues of MDS patients, suggesting an important role of PANoptosis in MDS pathogenesis. Caspase 8 (Casp8) is a master regulator of PANoptosis, which is downregulated in HSPCs from most MDS patients and abnormally spliced in HSPCs from MDS patients with SRSF2 mutation. To study the role of PANoptosis in hematopoiesis, we generated inducible Casp8 knockout mice (Casp8-/-). Mx1-Cre-Casp8-/- mice died of BM failure within 10 days of polyI:C injections due to depletion of HSPCs. Rosa-ERT2Cre-Casp8-/- mice are healthy without significant changes in BM hematopoiesis within the first 1.5 months after Casp8 deletion. Such mice developed BM failure upon infection or low dose polyI:C/LPS injections due to the hypersensitivity of Casp8-/- HSPCs to infection or inflammation-induced necroptosis which can be prevented by Ripk3 deletion. However, impaired self-renewal capacity of Casp8-/- HSPCs cannot be rescued by Ripk3 deletion due to activation of Ripk1-Tbk1 signaling. Most importantly, mice transplanted with Casp8-/- BM cells developed MDS-like disease within 4 months of transplantation as demonstrated by anemia, thrombocytopenia and myelodysplasia. Our study suggests an essential role for a balance in Casp8, Ripk3-Mlkl and Ripk1-Tbk1 activities in the regulation of survival and self-renewal of HSPCs, the disruption of which induces inflammation and BM failure, resulting in MDS-like disease.

Distinct roles of hematopoietic cytokines in the regulation of leukemia stem cells in murine MLL-AF9 leukemia.

Lymphoid-primed multipotent progenitor (LMPP)-like and granulocyte-monocyte progenitor (GMP)-like leukemia stem cells (LSCs) co-exist in the blood of most patients with acute myeloid leukemia (AML). Complete elimination of both types of LSCs is required to cure AML. Using an MLL-AF9-induced murine AML model, we studied the role of hematopoietic cytokines in the survival of LMPP- and GMP-like LSCs. We found that SCF or FLT3L promotes the survival of LMPP-like LSCs by stimulating Stat5-mediated Mcl1 expression, whereas interleukin-3 (IL-3) or IL-6 induces the survival of GMP-like LSCs by stimulating Stat3/nuclear factor κB (NF-κB)-mediated Bcl2 expression. Functional study demonstrated that, compared to AML cells cultured in IL-3 and IL-6 medium, AML cells in SCF- or Flt3L-only culture are highly clonogenic in in vitro culture and are highly leukemogenic in vivo. Our study suggests that co-inhibition of both STAT5-MCL1 and STAT3/NF-κB-BCL2 signaling might represent an improved treatment strategy against AML, specifically AML cases with a monocytic phenotype and/or FLT3 mutations.

Remarkably rapid, recent diversification of Cochemiea and Mammillaria in the Baja California, Mexico region.

PREMISE: The Cactaceae of northwestern Mexico and the southwestern United States constitute a major component of the angiosperm biodiversity of the region. The Mammilloid clade, (Cactaceae, tribe Cacteae), composed of the genera Cochemiea, Coryphantha, Cumarinia, Mammillaria, and Pelecyphora is especially species rich. We sought to understand the timing, geographical and climate influences correlated with expansion of the Mammilloid clade, through the Sonoran Desert into Baja California. METHODS: We reconstructed the historical biogeography of the Mammilloid clade, using Bayesian and maximum likelihood methods, based on a strongly supported molecular phylogeny. We also estimated divergence times, the timing of emergence of key characters, and diversification rates and rate shifts of the Mammilloid clade. RESULTS: We found that the most recent common ancestor of Cochemiea arrived in the Cape region of Baja California from the Sonoran Desert region approximately 5 million years ago, coinciding with the timing of peninsular rifting from the mainland, suggesting dispersal and vicariance as causes of species richness and endemism. The diversification rate for Cochemiea is estimated to be approximately 12 times that of the mean background diversification rate for angiosperms. Divergence time estimation shows that many of the extant taxa in Cochemiea and Baja California Mammillaria emerged from common ancestors 1 million to 200,000 years ago, having a mid-Pleistocene origin. CONCLUSIONS: Cochemiea and Mammillaria of the Baja California region are examples of recent, rapid diversification. Geological and climatic forces at multiple spatial and temporal scales are correlated with the western distributions of the Mammilloid clade.

Seed oil of Brucea javanica induces apoptosis through the PI3K/Akt signaling pathway in acute lymphocytic leukemia Jurkat cells.

Brucea javanica oil emulsion (BJOE) has been used to treat tumor in China for more than 40 years. However, its components and effectiveness in the treatment of acute lymphocytic leukemia (ALL) and its mechanism of anti-cancer activity remain unknown. In the current study, high-performance liquid chromatography-evaporative light scattering detector (HPLC-ELSD) was used to analyze the components of BJOE. Then, the anti-leukemia effects of BJOE were examined both in vitro and in vivo using ALL Jurkat cells and the p388 mouse leukemia transplant model, respectively. The primary ALL leukemia cells were also used to confirm the anti-leukemia effects of BJOE. The apoptotic-related results indicated that BJOE induced apoptosis in Jurkat cells and were suggestive of intrinsic apoptotic induction. Moreover, BJOE inhibited Akt (protein kinase B) activation and upregulated its downstream targets p53 and FoxO1 (forkhead box gene, group O-1) to initiate apoptosis. The activation of GSK3ß was also involved. Our findings demonstrate that BJOE has anti-leukemia effects on ALL cells and can induce apoptosis in Jurkat cells through the phosphoinositide3-kinase (PI3K) /Akt signaling pathway.

Projected climate change threatens significant range contraction of Cochemiea halei (Cactaceae), an island endemic, serpentine-adapted plant species at risk of extinction.

AIM: Threats faced by narrowly distributed endemic plant species in the face of the Earth's sixth mass extinction and climate change exposure are especially severe for taxa on islands. We investigated the current and projected distribution and range changes of Cochemiea halei, an endemic island cactus. This taxon is of conservation concern, currently listed as vulnerable on the International Union for the Conservation of Nature Red List and as a species of special concern under Mexican federal law. The goals of this study are to (a) identify the correlations between climate variables and current suitable habitat for C. halei; (b) determine whether the species is a serpentine endemic or has a facultative relationship with ultramafic soils; and (c) predict range changes of the species based on climate change scenarios. LOCATION: The island archipelago in Bahía Magdalena on the Pacific coast, Baja California Sur, Mexico. METHODS: We used temperature and precipitation variables at 30-arc second resolution and soil type, employing multiple species distribution modeling methods, to identify important climate and soil conditions driving current habitat suitability. The best model of current suitability is used to predict possible effects of four climate change scenarios based on best-case to worst-case representative concentration pathways, with projected climate data from two general circulation models, over two time periods. MAIN CONCLUSIONS: The occurrence of the species is found to be strongly correlated with ultramafic soils. The most important climate predictor for habitat suitability is annual temperature range. The species is predicted to undergo range contractions from 21% to 53%, depending on the severity and duration of exposure to climate change. The broader implications for a wide range of narrowly adapted, threatened, and endemic plant species indicate an urgent need for threat assessment based on habitat suitability and climate change modeling.

Molecular and cellular mechanisms of aging in hematopoietic stem cells and their niches.

Aging drives the genetic and epigenetic changes that result in a decline in hematopoietic stem cell (HSC) functioning. Such changes lead to aging-related hematopoietic/immune impairments and hematopoietic disorders. Understanding how such changes are initiated and how they progress will help in the development of medications that could improve the quality life for the elderly and to treat and possibly prevent aging-related hematopoietic diseases. Here, we review the most recent advances in research into HSC aging and discuss the role of HSC-intrinsic events, as well as those that relate to the aging bone marrow niche microenvironment in the overall processes of HSC aging. In addition, we discuss the potential mechanisms by which HSC aging is regulated.

ABL1, Overexpressed in Hepatocellular Carcinomas, Regulates Expression of NOTCH1 and Promotes Development of Liver Tumors in Mice.

BACKGROUND & AIMS: We investigated whether ABL proto-oncogene 1, non-receptor tyrosine kinase (ABL1) is involved in development of hepatocellular carcinoma (HCC). METHODS: We analyzed clinical and gene expression data from The Cancer Genome Atlas. Albumin-Cre (HepWT) mice and mice with hepatocyte-specific disruption of Abl1 (HepAbl-/- mice) were given hydrodynamic injections of plasmids encoding the Sleeping Beauty transposase and transposons with the MET gene and a catenin ß1 gene with an N-terminal truncation, which induces development of liver tumors. Some mice were then gavaged with the ABL1 inhibitor nilotinib or vehicle (control) daily for 4 weeks. We knocked down ABL1 with short hairpin RNAs in Hep3B and Huh7 HCC cells and analyzed their proliferation and growth as xenograft tumors in mice. We performed RNA sequencing and gene set enrichment analysis of tumors. We knocked down or overexpressed NOTCH1 and MYC in HCC cells and analyzed proliferation. We measured levels of phosphorylated ABL1, MYC, and NOTCH1 by immunohistochemical analysis of an HCC tissue microarray. RESULTS: HCC tissues had higher levels of ABL1 than non-tumor liver tissues, which correlated with shorter survival times of patients. HepWT mice with the MET and catenin ß1 transposons developed liver tumors and survived a median 64 days; HepAbl-/- mice with these transposons developed tumors that were 50% smaller and survived a median 81 days. Knockdown of ABL1 in human HCC cells reduced proliferation, growth as xenograft tumors in mice, and expression of MYC, which reduced expression of NOTCH1. Knockdown of NOTCH1 or MYC in HCC cells significantly reduced cell growth. NOTCH1 or MYC overexpression in human HCC cells promoted proliferation and rescued the phenotype caused by ABL1 knockdown. The level of phosphorylated (activated) ABL1 correlated with levels of MYC and NOTCH1 in human HCC specimens. Nilotinib decreased expression of MYC and NOTCH1 in HCC cell lines, reduced the growth of xenograft tumors in mice, and slowed growth of liver tumors in mice with MET and catenin ß1 transposons, reducing tumor levels of MYC and NOTCH1. CONCLUSIONS: HCC samples have increased levels of ABL1 compared with nontumor liver tissues, and increased levels of ABL1 correlate with shorter survival times of patients. Loss or inhibition of ABL1 reduces proliferation of HCC cells and slows growth of liver tumors in mice. Inhibitors of ABL1 might be used for treatment of HCC.

Myc-Miz1 signaling promotes self-renewal of leukemia stem cells by repressing Cebpα and Cebpδ.

c-Myc (Myc hereafter) is found to be deregulated and/or amplified in most acute myeloid leukemias (AMLs). Almost all AML cells are dependent upon Myc for their proliferation and survival. Thus, Myc has been proposed as a critical anti-AML target. Myc has Max-mediated transactivational and Myc-interacting zinc finger protein 1 (Miz1)-mediated transrepressional activities. The role of Myc-Max-mediated transactivation in the pathogenesis of AML has been well studied; however, the role of Myc-Miz1-mediated transrepression in AML is still somewhat obscure. Myc protein harboring a V394D mutation (MycV394D) is a mutant form of Myc that lacks transrepressional activity due to a defect in its ability to interact with Miz1. We found that, compared with Myc, the oncogenic function of MycV394D is significantly impaired. The AML/myeloproliferative disorder that develops in mice receiving MycV394D-transduced hematopoietic stem/progenitor cells (HSPCs) is significantly delayed compared with mice receiving Myc-transduced HSPCs. Using a murine MLL-AF9 AML model, we found that AML cells expressing MycV394D (intrinsic Myc deleted) are partially differentiated and show reductions in both colony-forming ability in vitro and leukemogenic capacity in vivo. The reduced frequency of leukemia stem cells (LSCs) among MycV394D-AML cells and their reduced leukemogenic capacity during serial transplantation suggest that Myc-Miz1 interaction is required for the self-renewal of LSCs. In addition, we found that MycV394D-AML cells are more sensitive to chemotherapy than are Myc-AML cells. Mechanistically, we found that Myc represses Miz1-mediated expression of CCAAT/enhancer-binding protein α (Cebpα) and Cebpδ, thus playing an important role in the pathogenesis of AML by maintaining the undifferentiated state and self-renewal capacity of LSCs.

In Vitro Expansion of Hematopoietic Stem Cells by Inhibition of Both GSK3 and p38 Signaling.

Hematopoietic stem cell (HSC) transplantation therapy is one of the most effective treatments for life-threatening hematopoietic diseases. Bone marrow (BM) and mobilized peripheral blood are the major sources of HSCs, but these resources are limited by a paucity of human leukocyte antigen (HLA)-matched donors. Umbilical cord blood (UCB) is the most promising alternative to obtain HSCs for transplantation therapy. However, UCB transplantation therapy is limited by low numbers of HSCs per unit of UCB. In vitro HSC expansion is believed to be the most effective and applicable strategy to address this issue. Here we report that a moderate concentration of GSK3 inhibitor promotes HSC expansion by inducing moderate levels of ß-catenin activity in HSCs. However, such a concentration of GSK3 inhibitor also stimulates myeloid cells to produce inflammatory cytokines, which attenuate HSC expansion by inducing p38 activation. Thus, when unpurified HSCs were used in culture, inhibition of p38-induced inflammatory cytokine signaling was required to ensure HSC expansion induced by the low concentration of GSK3 inhibitor. Our study suggests that the combination of a moderate concentration of p38 inhibitor plus a GSK3 inhibitor synergistically promotes the expansion of both murine BM HSCs and human UCB HSCs.

Focal Adhesion Kinase and ß-Catenin Cooperate to Induce Hepatocellular Carcinoma.

There is an urgent need to understand the molecular signaling pathways that drive or mediate the development of hepatocellular carcinoma (HCC). The focal adhesion kinase (FAK) gene protein tyrosine kinase 2 is amplified in 16.4% of The Cancer Genome Atlas HCC specimens, and its amplification leads to increased FAK mRNA expression. It is not known whether the overexpression of FAK alone is sufficient to induce HCC or whether it must cooperate in some ways with other oncogenes. In this study, we found that 34.8% of human HCC samples with FAK amplification also show ß-catenin mutations, suggesting a co-occurrence of FAK overexpression and ß-catenin mutations in HCC. We overexpressed FAK alone, constitutively active forms of ß-catenin (CAT) alone, or a combination of FAK and CAT in the livers of C57/BL6 mice. We found that overexpression of both FAK and CAT, but neither FAK nor CAT alone, in mouse livers was sufficient to lead to tumorigenesis. We further demonstrated that FAK's kinase activity is required for FAK/CAT-induced tumorigenesis. Furthermore, we performed RNA-sequencing analysis to identify the genes/signaling pathways regulated by FAK, CAT, or FAK/CAT. We found that FAK overexpression dramatically enhances binding of ß-catenin to the promoter of androgen receptor (AR), which leads to increased expression of AR in mouse livers. Moreover, ASC-J9, an AR degradation enhancer, suppressed FAK/CAT-induced HCC formation. Conclusion: FAK overexpression and ß-catenin mutations often co-occur in human HCC tissues. Co-overexpression of FAK and CAT leads to HCC formation in mice through increased expression of AR; this mouse model may be useful for further studies of the molecular mechanisms in the pathogenesis of HCC and could lead to the identification of therapeutic targets.

Inhibition of insulin-like growth factor 1 receptor enhances the efficacy of sorafenib in inhibiting hepatocellular carcinoma cell growth and survival.

Hepatocellular carcinoma (HCC) is the fifth most common primary cancer and second largest cause of cancer-related death worldwide. The first-line oral chemotherapeutic agent sorafenib only increases survival in patients with advanced HCC by less than 3 months. Most patients with advanced HCC have shown limited response rates and survival benefits with sorafenib. Although sorafenib is an inhibitor of multiple kinases, including serine/threonine-protein kinase c-Raf, serine/threonine-protein kinase B-Raf, vascular endothelial growth factor receptor (VEGFR)-1, VEGFR-2, VEGFR-3, and platelet-derived growth factor receptor ß, HCC cells are able to escape from sorafenib treatment using other pathways that the drug insufficiently inhibits. The aim of this study was to identify and target survival and proliferation pathways that enable HCC to escape the antitumor activity of sorafenib. We found that insulin-like growth factor 1 receptor (IGF1R) remains activated in HCC cells treated with sorafenib. Knockdown of IGF1R sensitizes HCC cells to sorafenib treatment and decreases protein kinase B (AKT) activation. Overexpression of constitutively activated AKT reverses the effect of knockdown of IGF1R in sensitizing HCC cells to treatment with sorafenib. Further, we found that ceritinib, a drug approved by the U.S. Food and Drug Administration for treatment of non-small cell lung cancer, effectively inhibits the IGF1R/AKT pathway and enhances the inhibitory efficacy of sorafenib in human HCC cell growth and survival in vitro, in a xenograft mouse model and in the c-Met/ß-catenin-driven HCC mouse model. Conclusion: Our study provides a biochemical basis for evaluation of a new combination treatment that includes IGF1R inhibitors, such as ceritinib and sorafenib, in patients with HCC. (Hepatology Communications 2018;2:732-746).

Caspase-3 suppresses diethylnitrosamine-induced hepatocyte death, compensatory proliferation and hepatocarcinogenesis through inhibiting p38 activation.

It is critical to understand the molecular mechanisms of hepatocarcinogenesis in order to prevent or treat hepatocellular carcinoma (HCC). The development of HCC is commonly associated with hepatocyte death and compensatory proliferation. However, the role of Caspase-3, a key apoptotic executor, in hepatocarcinogenesis is unknown. In this study, we used Caspase-3-deficient mice to examine the role of Caspase-3 in hepatocarcinogenesis in a chemical (diethylnitrosamine, DEN)-induced HCC model. We found that Caspase-3 deficiency significantly increased DEN-induced HCC. Unexpectedly, Caspase-3 deficiency increased apoptosis induced by DEN and the subsequent compensatory proliferation. Intriguingly, we discovered that Caspase-3 deficiency increased the activation of p38 with and without DEN treatment. Moreover, we demonstrated that TNFα and IL1α stimulated increased activation of p38 in Caspase-3 KO hepatocytes compared with wild-type hepatocytes. Finally, we found that inhibition of p38 by SB202190 abrogated enhanced hepatocyte death, compensatory proliferation and HCC induced by DEN in Caspase-3-deficient mice. Overall, our data suggest that Caspase-3 inhibits chemical-induced hepatocarcinogenesis by suppressing p38 activation and hepatocyte death.

Sensitizing leukemia stem cells to NF-κB inhibitor treatment in vivo by inactivation of both TNF and IL-1 signaling.

We previously reported that autocrine TNF-α (TNF) is responsible for JNK pathway activation in a subset of acute myeloid leukemia (AML) patient samples, providing a survival/proliferation signaling parallel to NF-κB in AML stem cells (LSCs). In this study, we report that most TNF-expressing AML cells (LCs) also express another pro-inflammatory cytokine, IL1ß, which acts in a parallel manner. TNF was produced primarily by LSCs and leukemic progenitors (LPs), whereas IL1ß was mainly produced by partially differentiated leukemic blasts (LBs). IL1ß also stimulates an NF-κB-independent pro-survival and proliferation signal through activation of the JNK pathway. We determined that co-inhibition of signaling stimulated by both TNF and IL1ß synergizes with NF-κB inhibition in eliminating LSCs both ex vivo and in vivo. Our studies show that such treatments are most effective in M4/5 subtypes of AML.

Necroptosis in spontaneously-mutated hematopoietic cells induces autoimmune bone marrow failure in mice.

Acquired aplastic anemia is an autoimmune-mediated bone marrow failure syndrome. The mechanism by which such an autoimmune reaction is initiated is unknown. Whether and how the genetic lesions detected in patients cause autoimmune bone marrow failure have not yet been determined. We found that mice with spontaneous deletion of the TGFß-activated kinase-1 gene in a small subset of hematopoietic cells developed bone marrow failure which resembled the clinical manifestations of acquired aplastic anemia patients. Bone marrow failure in such mice could be reversed by depletion of CD4+ T lymphocytes or blocked by knockout of interferon-γ, suggesting a Th1-cell-mediated autoimmune mechanism. The onset and progression of bone marrow failure in such mice were significantly accelerated by the inactivation of tumor necrosis factor-α signaling. Tumor necrosis factor-α restricts autoimmune bone marrow failure by inhibiting type-1 T-cell responses and maintaining the function of myeloid-derived suppressor cells. Furthermore, we determined that necroptosis among a small subset of mutant hematopoietic cells is the cause of autoimmune bone marrow failure because such bone marrow failure can be prevented by deletion of receptor interacting protein kinase-3 Our study suggests a novel mechanism to explain the pathogenesis of autoimmune bone marrow failure.

FAK deletion accelerates liver regeneration after two-thirds partial hepatectomy.

Understanding the molecular mechanisms of liver regeneration is essential to improve the survival rate of patients after surgical resection of large amounts of liver tissue. Focal adhesion kinase (FAK) regulates different cellular functions, including cell survival, proliferation and cell migration. The role of FAK in liver regeneration remains unknown. In this study, we found that Fak is activated and induced during liver regeneration after two-thirds partial hepatectomy (PHx). We used mice with liver-specific deletion of Fak and investigated the role of Fak in liver regeneration in 2/3 PHx model (removal of 2/3 of the liver). We found that specific deletion of Fak accelerates liver regeneration. Fak deletion enhances hepatocyte proliferation prior to day 3 post-PHx but attenuates hepatocyte proliferation 3 days after PHx. Moreover, we demonstrated that the deletion of Fak in liver transiently increases EGFR activation by regulating the TNFα/HB-EGF axis during liver regeneration. Furthermore, we found more apoptosis in Fak-deficient mouse livers compared to WT mouse livers after PHx. CONCLUSION: Our data suggest that Fak is involved in the process of liver regeneration, and inhibition of FAK may be a promising strategy to accelerate liver regeneration in recipients after liver transplantation.

FAK Kinase Activity Is Required for the Progression of c-MET/ß-Catenin-Driven Hepataocellular Carcinoma.

There is an urgent need to develop new and more effective therapeutic strategies and agents to treat hepatocellular carcinoma (HCC). We have recently found that deletion of Fak in hepatocytes before tumors form inhibits tumor development and prolongs survival of animals in a c-MET (MET)/ß-catenin (CAT)-driven HCC mouse model. However, it has yet to be determined whether FAK expression in hepatocytes promotes MET/CAT-induced HCC progression after tumor initiation. In addition, it remains unclear whether FAK promotes HCC development through its kinase activity. We generated hepatocyte-specific inducible Fak-deficient mice (Alb-creERT2; Fak(flox/flox)) to examine the role of FAK in HCC progression. We reexpressed wild-type and mutant FAK in Fak-deficient mice to determine FAK's kinase activity in HCC development. We also examined the efficacy of a FAK kinase inhibitor PF-562271 on HCC inhibition. We found that deletion of Fak after tumors form significantly repressed MET/CAT-induced tumor progression. Ectopic FAK expression restored HCC formation in hepatocyte-specific Fak-deficient mice. However, overexpression of a FAK kinase-dead mutant led to reduced tumor load compared to mice that express wild-type FAK. Furthermore, PF-562271 significantly suppressed progression of MET/CAT-induced HCC. Fak kinase activity is important for MET/CAT-induced HCC progression. Inhibiting FAK kinase activity provides a potential therapeutic strategy to treat HCC.

Hematopoietic Stem Cell Activity Is Regulated by Pten Phosphorylation Through a Niche-Dependent Mechanism.

The phosphorylated form of Pten (p-Pten) is highly expressed in >70% of acute myeloid leukemia samples. However, the role of p-Pten in normal and abnormal hematopoiesis has not been studied. We found that Pten protein levels are comparable among long-term (LT) hematopoietic stem cells (HSCs), short-term (ST) HSCs, and multipotent progenitors (MPPs); however, the levels of p-Pten are elevated during the HSC-to-MPP transition. To study whether p-Pten is involved in regulating self-renewal and differentiation in HSCs, we compared the effects of overexpression of p-Pten and nonphosphorylated Pten (non-p-Pten) on the hematopoietic reconstitutive capacity (HRC) of HSCs. We found that overexpression of non-p-Pten enhances the LT-HRC of HSCs, whereas overexpression of p-Pten promotes myeloid differentiation and compromises the LT-HRC of HSCs. Such phosphorylation-regulated Pten functioning is mediated by repressing the cell:cell contact-induced activation of Fak/p38 signaling independent of Pten's lipid phosphatase activity because both p-Pten and non-p-Pten have comparable activity in repressing PI3K/Akt signaling. Our studies suggest that, in addition to repressing PI3K/Akt/mTor signaling, non-p-Pten maintains HSCs in bone marrow niches via a cell-contact inhibitory mechanism by inhibiting Fak/p38 signaling-mediated proliferation and differentiation. In contrast, p-Pten promotes the proliferation and differentiation of HSCs by enhancing the cell contact-dependent activation of Src/Fak/p38 signaling. Stem Cells 2016;34:2130-2144.

Inhibition of SIRT2 suppresses hepatic fibrosis.

Liver fibrosis can progress to cirrhosis and result in serious complications of liver disease. The pathogenesis of liver fibrosis involves the activation of hepatic stellate cells (HSCs), the underlying mechanisms of which are not fully known. Emerging evidence suggests that the classic histone deacetylases play a role in liver fibrosis, but the role of another subfamily of histone deacetylases, the sirtuins, in the development of hepatic fibrosis remains unknown. In this study, we found that blocking the activity of sirtuin 2 (SIRT2) by using inhibitors or shRNAs significantly suppressed fibrogenic gene expression in HSCs. We further demonstrated that inhibition of SIRT2 results in the degradation of c-MYC, which is important for HSC activation. In addition, we discovered that inhibition of SIRT2 suppresses the phosphorylation of ERK, which is critical for the stabilization of c-MYC. Moreover, we found that Sirt2 deficiency attenuates the hepatic fibrosis induced by carbon tetrachloride (CCl4) and thioacetamide (TAA). Furthermore, we showed that SIRT2, p-ERK, and c-MYC proteins are all overexpressed in human hepatic fibrotic tissues. These data suggest a critical role for the SIRT2/ERK/c-MYC axis in promoting hepatic fibrogenesis. Inhibition of the SIRT2/ERK/c-MYC axis represents a novel strategy to prevent and to potentially treat liver fibrosis and cirrhosis.

FAK mediates a compensatory survival signal parallel to PI3K-AKT in PTEN-null T-ALL cells.

Mutations and inactivation of phosphatase and tensin homolog deleted from chromosome 10 (PTEN) are observed in 15%-25% of cases of human T cell acute lymphoblastic leukemia (T-ALL). Pten deletion induces myeloproliferative disorders (MPDs), acute myeloid leukemia (AML), and/or T-ALL in mice. Previous studies attributed Pten-loss-related hematopoietic defects and leukemogenesis to excessive activation of phosphatidylinositol 3-kinase (PI3K)/AKT/mTOR signaling. Although inhibition of this signal dramatically suppresses the growth of PTEN-null T-ALL cells in vitro, treatment with inhibitors of this pathway does not cause a complete remission in vivo. Here, we report that focal adhesion kinase (Fak), a protein substrate of Pten, also contributes to T-ALL development in Pten-null mice. Inactivation of the FAK signaling pathway by either genetic or pharmacologic methods significantly sensitizes both murine and human PTEN-null T-ALL cells to PI3K/AKT/mTOR inhibition when cultured in vitro on feeder layer cells or a matrix and in vivo.

The synergistic repressive effect of NF-κB and JNK inhibitor on the clonogenic capacity of Jurkat leukemia cells.

Deregulation of Nuclear Transcription Factor-κB (NF-κB) and Jun N-terminal kinase (JNK) signaling is commonly detected in leukemia, suggesting an important role for these two signaling pathways in the pathogenesis of leukemia. In this study, using Jurkat cells, an acute T-lymphoblastic leukemia (T-ALL) cell line, we evaluated the effects of an NF-κB inhibitor and a JNK inhibitor individually and in combination on the proliferation, survival and clonogenic capacity of leukemic cells. We found that leukemic stem/progenitor cells (LSPCs) were more sensitive to NF-κB inhibitor treatment than were healthy hematopoietic stem/progenitor cells (HSPCs), as shown by a reduction in the clonogenic capacity of the former. Inactivation of NF-κB leads to the activation of JNK signaling in both leukemic cells and healthy HSPCs. Interestingly, JNK inhibitor treatment enhanced the repressive effects of NF-κB inhibitor on LSPCs but prevented such repression in HSPCs. Our data suggest that JNK signaling stimulates proliferation/survival in LSPCs but is a death signal in HSPCs. The combination of NF-κB inhibitor and JNK inhibitor might provide a better treatment for T-ALL leukemia by synergistically killing LSPCs while simultaneously preventing the death of normal HPCs.