RAS oncogene signal strength regulates matrisomal gene expression and tumorigenicity of mouse keratinocytes.

Environmental and molecular carcinogenesis are linked by the discovery that chemical carcinogen induced-mutations in the Hras or Kras genes drives tumor development in mouse skin. Importantly, enhanced expression or allele amplification of the mutant Ras gene contributes to selection of initiated cells, tumor persistence, and progression. To explore the consequences of Ras oncogene signal strength, primary keratinocytes were isolated and cultured from the LSL-HrasG12D and LSL-KrasG12D C57BL/6J mouse models and the mutant allele was activated by adeno-Cre recombinase. Keratinocytes expressing one (H) or two (HH) mutant alleles of HrasG12D, one KrasG12D allele (K), or one of each (HK) were studied. All combinations of activated Ras alleles stimulated proliferation and drove transformation marker expression, but only HH and HK formed tumors. HH, HK, and K sustained long-term keratinocyte growth in vitro, while H and WT could not. RNA-Seq yielded two distinct gene expression profiles; HH, HK, and K formed one cluster while H clustered with WT. Weak MAPK activation was seen in H keratinocytes but treatment with a BRAF inhibitor enhanced MAPK signaling and facilitated tumor formation. K keratinocytes became tumorigenic when they were isolated from mice where the LSL-KrasG12D allele was backcrossed from the C57BL/6 onto the FVB/N background. All tumorigenic keratinocytes but not the non-tumorigenic precursors shared a common remodeling of matrisomal gene expression that is associated with tumor formation. Thus, RAS oncogene signal strength determines cell-autonomous changes in initiated cells that are critical for their tumor-forming potential.

Pseudophosphatase MK-STYX Alters Histone Deacetylase 6 Cytoplasmic Localization, Decreases Its Phosphorylation, and Increases Detyrosination of Tubulin.

The catalytically inactive mitogen-activated protein (MAP) kinase phosphatase, MK-STYX (MAPK (mitogen-activated protein kinase) phosphoserine/threonine/tyrosine-binding protein) interacts with the stress granule nucleator G3BP-1 (Ras-GAP (GTPase-activating protein) SH3 (Src homology 3) domain-binding protein-1), and decreases stress granule (stalled mRNA) formation. Histone deacetylase isoform 6 (HDAC6) also binds G3BP-1 and serves as a major component of stress granules. The discovery that MK-STYX and HDAC6 both interact with G3BP-1 led us to investigate the effects of MK-STYX on HDAC6 dynamics. In control HEK/293 cells, HDAC6 was cytosolic, as expected, and formed aggregates under conditions of stress. In contrast, in cells overexpressing MK-STYX, HDAC6 was both nuclear and cytosolic and the number of stress-induced aggregates significantly decreased. Immunoblots showed that MK-STYX decreases HDAC6 serine phosphorylation, protein tyrosine phosphorylation, and lysine acetylation. HDAC6 is known to regulate microtubule dynamics to form aggregates. MK-STYX did not affect the organization of microtubules, but did affect their post-translational modification. Tubulin acetylation was increased in the presence of MK-STYX. In addition, the detyrosination of tubulin was significantly increased in the presence of MK-STYX. These findings show that MK-STYX decreases the number of HDAC6-containing aggregates and alters their localization, sustains microtubule acetylation, and increases detyrosination of microtubules, implicating MK-STYX as a signaling molecule in HDAC6 activity.

Safety of postdischarge extended venous thromboembolism prophylaxis after hepatopancreatobiliary surgery.

BACKGROUND: The risk of venous thromboembolism (VTE) after hepatopancreatobiliary (HPB) surgery is high. Extended postdischarge prophylaxis in this patient population has been controversial. This study aimed to examine the safety of postdischarge extended VTE prophylaxis in patients at high risk of VTE events after HPB surgery. METHODS: Adult patients risk stratified as very high risk of VTE who underwent HPB operations between 2014 and 2020 at a quaternary care center were included. Patients were matched 1:2 extended VTE prophylaxis to the control group (patients who did not receive extended prophylaxis). Analyses compared the proportions of adverse bleeding events between groups. RESULTS: A total of 307 patients were included: 103 in the extended prophylaxis group and 204 in the matched control group. Demographics were similar between groups. More patients in the extended VTE prophylaxis group had a history of VTE (9% vs 3%; P = .045). There was no difference in bleeding events between the extended VTE prophylaxis and the control group (6% vs 2%; P = .091). Of the 6 patients with bleeding events in the VTE prophylaxis group, 5 had gastrointestinal (GI) bleeding, and 1 had hemarthrosis. Of the 4 patients with bleeding events in the control group, 1 had intra-abdominal bleeding, 2 had GI bleeding, and 1 had intra-abdominal and GI bleeding. CONCLUSION: Patients discharged with extended VTE prophylaxis after HPB surgery did not experience more adverse bleeding events compared with a matched control group. Routine postdischarge extended VTE prophylaxis is safe in patients at high risk of postoperative VTE after HPB surgery.