Non-canonical Small GTPase RBJ Promotes NSCLC Progression Through the Canonical MEK/ERK Signaling Pathway.

BACKGROUND: Although the majority of members belonging to the small GTPase Ras superfamily have been studied in several malignancies, the function of RBJ has remained unclear, particularly in non-small cell lung cancer (NSCLC). OBJECTIVE: The research aims to determine the function of RBJ in NSCLC. METHODS: The levels of RBJ protein in tumor tissue and para-carcinoma normal tissue were ascertained via immunohistochemistry (IHC). The growth, migration, and invasion of NSCLC cells were assessed by 5- ethynyl-2-deoxyuridine (EdU) assay, colony formation, cell counting kit-8 (CCK8), transwell and wound healing assays. Furthermore, a nude mouse xenograft model was established to study the function of RBJ in tumorigenesis in vivo. RESULTS: The IHC analysis revealed that the protein levels of RBJ were notably increased in tumor tissue and positively associated with the clinical stage. In addition, the knockdown of RBJ restrained the growth, invasion, and migration of NSCLC cell lines by inhibiting the epithelial-mesenchymal transition (EMT) through the MEK/ERK signaling pathway. Accordingly, opposite results were observed when RBJ was overexpressed. In addition, the overexpression of RBJ accelerated tumor formation by A549 cells in nude mice. CONCLUSION: RBJ promoted cancer progression in NSCLC by activating EMT via the MEK/ERK signaling. Thus, RBJ could be used as a potential therapeutic against NSCLC.

Bronchial smooth muscle cell in asthma: where does it fit?

Asthma is a frequent respiratory condition whose pathophysiology relies on altered interactions between bronchial epithelium, smooth muscle cells (SMC) and immune responses. Those leads to classical hallmarks of asthma: airway hyper-responsiveness, bronchial remodelling and chronic inflammation. Airway smooth muscle biology and pathophysiological implication in asthma are now better understood. Precise deciphering of intracellular signalling pathways regulating smooth muscle contraction highlighted the critical roles played by small GTPases of Rho superfamily. Beyond contractile considerations, active involvement of airway smooth muscle in bronchial remodelling mechanisms is now established. Not only cytokines and growth factors, such as fibroblats growth factor or transforming growth factor-ß, but also extracellular matrix composition have been demonstrated as potent phenotype modifiers for airway SMC. Although basic science knowledge has grown significantly, little of it has translated into improvement in asthma clinical practice. Evaluation of airway smooth muscle function is still limited to its contractile activity. Moreover, it relies on tools, such as spirometry, that give only an overall assessment and not a specific one. Interesting technics such as forced oscillometry or specific imagery (CT and MRI) give new perspectives to evaluate other aspects of airway muscle such as bronchial remodelling. Finally, except for the refinement of conventional bronchodilators, no new drug therapy directly targeting airway smooth muscle proved its efficacy. Bronchial thermoplasty is an innovative and efficient therapeutic strategy but is only restricted to a small proportion of severe asthmatic patients. New diagnostic and therapeutic strategies specifically oriented toward airway smooth muscle are needed to improve global asthma care.

Farnesyl diphosphate synthase regulated endothelial proliferation and autophagy during rat pulmonary arterial hypertension induced by monocrotaline.

BACKGROUND: The proliferation ability and autophagy level of pulmonary artery endothelial cells (PAECs) play an important role in promoting the development of pulmonary artery hypertension (PAH), and there is still no effective treatment for PAH. Farnesyl diphosphate synthase (FDPS) is a key enzyme in the mevalonate pathway. The intermediate metabolites of this pathway are closely related to the activity of autophagy-associated small G proteins, including Ras-related C3 botulinum toxin substrate 1 (Rac1). Studies have shown that the mevalonate pathway affects the activation levels of different small G proteins, autophagy signaling pathways, vascular endothelial function, and so on. However, the exact relationship between them is still unclear in PAH. METHOD: In vitro, western blotting and mRFP-GFP-LC3 puncta formation assays were used to observe the expression of FDPS and the level of autophagy in PAECs treated with monocrotaline pyrrole (MCTP). In addition, cell proliferation and migration assays were used to assess the effect of FDPS on endothelial function, and Rac1 activity assays were used to evaluate the effect of Rac1 activation on PAEC autophagy via the PI3K/AKT/mTOR signaling pathway. In vivo, the right heart catheterization method, hematoxylin and eosin (H&E) staining and western blotting were used to determine the effect of FDPS on PAEC autophagy and monocrotaline (MCT)-induced PAH. RESULTS: We show that the expression of FDPS is increased in the PAH module in vitro and in vivo, concomitant with the induction of autophagy and the activation of Rac1. Our data demonstrate that inhibition of FDPS ameliorates endothelial function and decreases MCT-induced autophagy levels. Mechanistically, we found that FDPS promotes autophagy, Rac1 activity and endothelial disfunction through the PI3K/AKT/mTOR signaling pathway. CONCLUSION: Our study suggests that FDPS contributes to active small G protein-induced autophagy during MCT-induced PAH, which may serve as a potential therapeutic target against PAH.

RAB11A Expression Is Associated With Cancer Aggressiveness Through Regulation of FGFR-Signaling in Lung Squamous Cell Carcinoma.

BACKGROUND: Fibroblast growth factor receptor (FGFR)-signaling in lung squamous cell carcinoma (LSCC) is associated with cancer aggressiveness and poor prognosis. Small GTPase RAB11A regulates the recycling of membrane proteins such as FGFR. This study evaluated the potential of RAB11A as a new therapeutic target for LSCC through its regulation of FGFR-signaling. METHODS: Immunohistochemical analysis of 84 LSCC samples was performed to determine the correlation between RAB11A expression, clinicopathologic features, and prognosis. Alterations in FGFR-signaling were assessed in RAB11A-suppressed and RAB11A-overexpressed LSCC cells both in vitro and in vivo. RESULTS: The study identified RAB11A as a strong predictor of poor prognosis in the LSCC cohort. Cell proliferation and invasion were promoted and inhibited respectively in RAB11A-overexpressed and RAB11A -suppressed LSCC cells. In RAB11A-overexpressed and RAB11A-suppressed LSCC cells, FGFR-signaling was respectively up- and downregulated. The viability of the cells treated with nintedanib and lenvatinib was greater in RAB11A-overexpressing cells than in control cells. The in vivo tumor growth and micro-vessel density of RAB11A-overexpressing tumors were significantly higher than in the control cells. CONCLUSION: As a potentially valuable prognostic marker, RAB11A is a promising therapeutic target for LSCC. Evaluation of RAB11A may be useful for identification of LSCC in patients whose cancer is refractory to FGFR inhibitors.

Overexpressed or hyperactivated Rac1 as a target to treat hepatocellular carcinoma.

Despite novel targeted and immunotherapies, the prognosis remains bleak for patients with hepatocellular carcinoma (HCC), especially for advanced and/or metastatic forms. The rapid emergence of drug resistance is a major obstacle in the success of chemo-, targeted-, immuno-therapies of HCC. Novel targets are needed. The prominent roles of the small GTPase Rac1 in the development and progression of HCC are discussed here, together with its multiple protein partners, and the targeting of Rac1 with RNA-based regulators and small molecules. We discuss the oncogenic functions of Rac1 in HCC, including the contribution of Rac1 mutants and isoform Rac1b. Rac1 is a ubiquitous target, but the protein is frequently overexpressed and hyperactivated in HCC. It contributes to the aggressivity of the disease, with key roles in cancer cell proliferation, tumor metastasis and resistance to treatment. Small molecule targeting Rac1, indirectly or directly, have shown anticancer effects in HCC experimental models. Rac1-binding agents such as EHT 1864 and analogues offer novel opportunities to combat HCC. We discuss the different modalities to repress Rac1 overactivation in HCC with small molecules and the combination with reference drugs to promote cancer cell death and to repress cell invasion. We highlight the necessity to combine Rac1-targeted approach with appropriate biomarkers to select Rac1 activated tumors. Our analysis underlines the prominent oncogenic functions of Rac1 in HCC and discuss the modalities to target this small GTPase. Rac1 shall be considered as a valid target to limit the acquired and intrinsic resistance of HCC tumors and their metastatic potential.

Euphorbia factor L3 ameliorates rheumatoid arthritis by suppressing the inflammatory response by targeting Rac family small GTPase 1.

Euphorbia factor L3 (EFL3) is extracted from Euphorbia lathyris and is known for its anti-inflammatory properties. This study focused on the potential anti-inflammatory and therapeutic effects of EFL3 on rheumatoid arthritis (RA) using fibroblast-like synoviocytes (FLSs) and arthritis animal models. Functional analysis showed that EFL3 could ameliorate the inflammatory phenotype of FLSs derived from RA patients, as evidenced by the decreases in cell viability, migration, invasion and cytokine production. Luciferase activity, Western blotting and immunofluorescence assays demonstrated that EFL3 inhibited the nuclear translocation of the p65 subunit and the subsequent activation of the nuclear factor kappa-Β (NF-κB) pathway. Furthermore, the therapeutic effects of EFL3 against arthritic progression were evidenced by decreases in joint swelling, arthritis scores, inflammatory factor production, synovial hyperplasia, and bone destruction in collagen-induced arthritis (CIA) and tumor necrosis factor-α (TNF-α) transgenic (TNF-tg) mouse models. Molecular analysis identified Rac family small GTPase 1 (Rac1) as the potential target that was required for EFL3-mediated suppression of the inflammatory RA FLS phenotype. In summary, this study uncovered the therapeutic potential of EFL3 in RA, which suggests its future clinical use.

Understanding Plastid Vesicle Transport - Could it Provide Benefit for Human Medicine?

BACKGROUND: In plants, vesicle transport occurs in the secretory pathway in the cytosol, between the membranes of different compartments. Several protein components have been identified to be involved in the process and their functions were characterized. Both cargos and other molecules (such as hormones) have been shown to use vesicle transport, although the major constituents of vesicles are lipids which are transferred from donor to acceptor membranes. In humans, malfunction of the cytosolic vesicle transport system leads to different diseases. METHOD: To better understand and ultimately cure these human diseases, studying other model systems such as yeast can be beneficial. Plants with their cytosolic vesicle transport system could serve as another model system. However, this review focuses on plant vesicles not present in the cytosol but in the chloroplasts, where lipids produced in the surrounding envelope are transported through the aqueous stroma to the thylakoid membranes. Although chloroplast vesicles have found both biochemical and ultrastructural support, only two proteins have been characterized as components of the pathway. However, using bioinformatics a number of other proteins have been suggested as homologs to the cytosolic system. RESULTS & CONCLUSION: Based on these findings vesicles of chloroplasts are likely most similar to the vesicles trafficking from ER to Golgi, or may even be unique, but important experimental support is yet lacking. In this review, proposed vesicle transport components in chloroplasts are presented, and their possible future implementation for human medicine is discussed.

Induction of GDNF and BDNF by hRheb(S16H) transduction of SNpc neurons: neuroprotective mechanisms of hRheb(S16H) in a model of Parkinson's disease.

The transduction of dopaminergic (DA) neurons with human ras homolog enriched in brain, which has a S16H mutation [hRheb(S16H)] protects the nigrostriatal DA projection in the 6-hydroxydopamine (6-OHDA)-treated animal model of Parkinson's disease (PD). However, it is still unclear whether the expression of active hRheb induces the production of neurotrophic factors such as glial cell line-derived neurotrophic factor (GDNF) and brain-derived neurotrophic factor (BDNF), which are involved in neuroprotection, in mature neurons. Here, we show that transduction of nigral DA neurons with hRheb(S16H) significantly increases the levels of phospho-cyclic adenosine monophosphate (cAMP) response element-binding protein (p-CREB), GDNF, and BDNF in neurons, which are attenuated by rapamycin, a specific inhibitor of mammalian target of rapamycin complex 1 (mTORC1). Moreover, treatment with specific neutralizing antibodies for GDNF and BDNF reduced the protective effects of hRheb(S16H) against 1-methyl-4-phenylpyridinium (MPP(+))-induced neurotoxicity. These results show that activation of hRheb/mTORC1 signaling pathway could impart to DA neurons the important ability to continuously produce GDNF and BDNF as therapeutic agents against PD.

Levels of specific antibodies towards the major antigenic determinant of slime-producing Staphylococcus epidermidis determined by an enzyme immunoassay and their protective effect in experimental keratitis.

Staphylococcus epidermidis is an important cause of bacterial keratitis. Certain S. epidermidis strains produce an extracellular slime layer rich in an acidic polysaccharide with a molecular size of 20 kDa (20-kDa PS). We have demonstrated that the level of 20-kDa PS-specific antibodies significantly rises after establishment of slime-producing S. epidermidis bacteraemia and, furthermore, that rabbit polyclonal antibodies to 20-kDa PS opsonize cells of slime-producing S. epidermidis to a great degree and promote their clearance by polymorphonuclear cells (Arch. Biochem. Biophys. 342 (1997) 389; J. Pharm. Biomed. Anal. 22 (2000) 1029). The purpose of this study was to examine the protective and therapeutic effects both of active immunization, using 20-kDa PS as antigen, and of passive administration of specific antibodies towards the 20-kDa PS in a rabbit keratitis model. For active immunization, 20 rabbits were subcutaneously immunized with 20-kDa PS, whereas for passive immunization specific polyclonal IgG antibodies against 20-kDa PS were administered to 20 rabbits 1 day before induction of infection. Clinical observations were made weekly for 1 month and levels of 20-kDa PS antibodies in serum and aqueous humor in both immunization groups were determined by an enzyme immunoassay. The levels of specific anti-20-kDa PS IgG in serum and aqueous humor following either active or passive immunization were significantly higher as compared with control groups (P<0.001). Although, actively immunized rabbits showed significantly less corneal damage than control animals, passively immunized ones were significantly better protected as compared with both control and those actively immunized. Obtained results suggest that 20-kDa PS plays crucial role in the pathogenesis of S. epidermidis keratitis and that both types of immunization significantly protect against corneal S. epidermidis pathology and damage.

[Effect of nm23H1 on proliferation, tumor formation and metastasis of hepatocarcinoma].

OBJECTIVE: To study the effects of nm23H1 on in vitro proliferation, in vivo tumor formation and metastasis of hepatocarcinoma. METHODS: Vectors containing insert of either sense or antisense nm23H1 cDNA under the albumin enhancer/promoter for liver cell-specific expression were constructed. The expression vectors were transfected into hepatocarcinoma cell line SMMC-7721. Stable cell clones with maximum and minimum expression of nm23 were obtained. Cell growth was measured in vitro and metastasis was observed in vivo by injecting cells into nude mice, subcutaneously or under the spleen capsule. RESULTS: In vitro, cell clone expressing antisense nm23H1 grew faster than that expressing sense nm23H1. In nude mice, the subcutaneously inoculated tumor grew faster and more aggressively in mice bearing antisense nm23H1 transfected hepatoma. So did metastasis of the antisense nm23H1 transfected hepatoma under the spleen capsule. CONCLUSION: Apart from suppressing metastasis, nm23H1 significantly suppresses tumor cell proliferation in vitro and tumor growth in vivo.