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As in other mammalian tissues, the extracellular matrix (ECM) of skin functions as mechanical support and regulative environment that guides the behavior of the cells. ECM is a gel-like structure that is primarily composed of structural and nonstructural proteins. While the content of structural proteins is stable, the level of nonstructural ECM proteins, such as thrombospondin-4 (THBS4), is dynamically regulated. In a previous work we demonstrated that THBS4 stimulated cutaneous wound healing. In this work we discovered that in addition to proliferation, THBS4 stimulated the migration of primary keratinocytes in 3D. By using a proteotransciptomic approach we found that stimulation of keratinocytes with THBS4 regulated the activity of signaling pathways linked to proliferation, migration, inflammation and differentiation. Interestingly, some of the regulated genes (eg IL37, TSLP) have been associated with the pathogenesis of atopic dermatitis (AD). We concluded that THBS4 is a promising candidate for novel wound healing therapies and suggest that there is a potential convergence of pathways that stimulate cutaneous wound healing with those active in the pathogenesis of inflammatory skin diseases. • THBS4 activates key pathways involved in keratinocyte migration, proliferation and differentiation.. • THBS4 activated intracellular signalling pathways in keratinocytes contribute to the cutaneous wound healing. • Pathways activated by THBS4 in primary keratinocytes overlap with those involved in the pathogenesis of AD. [ABSTRACT FROM AUTHOR]

Human mesenchymal stromal cells (MSC) are an important tool for basic and translational research. Large amounts of MSC are required for in vitro and in vivo studies, however, the limited life-span and differentiation ability in vitro hamper their optimal use. Here we report that 1:1 mixture of L15 and mTeSR1 culture media increased the life-span of IPI-SA3-C4, a normal non-immortalized human subcutaneous preadipocyte strain by 20% while retaining their adipogenic capacity and stable karyotype. The increased proliferative capacity was accompanied by increased expression of the stem markers POU5F1 , SOX2 , MYC and hTERT , and inhibition of hTERT activity abolished the growth advantage of L15-mTeSR1. Consequently, the described MSC culture would considerably enhance the utility of MSC for in vitro studies. [ABSTRACT FROM AUTHOR]

The lack of primary liver tumor cells has hampered testing of potential chemotherapeutic agents in vitro. To overcome this issue we developed a primary mouse liver tumor cell line K07074. The K07074 cells were immortal, exhibited a biliary phenotype, formed colonies in soft agar and displayed an increase in Hedgehog, Notch and Akt signaling. To study the effect of single and combined inhibition of the liver tumor-related pathways on the growth of K07074 cells we treated these with small-molecule antitumor agents. While the inhibition of Akt and Notch pathways strongly inhibited the growth of K07074 cells the inhibition of Wnt and Hedgehog pathways was less efficient in cell growth suppression. Interestingly, the inhibition of Akt pathway at the level of Akt-Pdpk1 interaction was sufficient to suppress the growth of tumor cells and no significant additive effect could be detected when co-treated with the inhibitors of Wnt, Hedgehog or Notch pathways. Only when suboptimal doses of Akt-Pdpk1 interaction inhibitor NSC156529 were used an additive effect with Notch inhibition was seen. We conclude that the Akt pathway inhibitor NSC156529 is potentially useful as single treatment for liver tumors with hyperactivated Akt signaling. [ABSTRACT FROM AUTHOR]

Olfactomedin 4 (OLFM4), a multifunctional matricellular protein, is involved in regulation of angiogenesis, innate immunity, inflammation, tumorigenesis and metastasis formation via modulation of important cellular processes like adhesion, proliferation, differentiation as well as apoptosis. In our previous work we demonstrated the upregulation of OLFM4 during liver regeneration and cutaneous wound healing. Here we studied the outcomes of OLFM4 downregulation in human immortalized keratinocytes – the HaCaT cells. The suppression of OLFM4 inhibited migration but enhanced the proliferation of these cells. By using proteomic, and phosphoproteomic analysis, we found that OLFM4 downregulation induced changes in the levels of 184 proteins and 348 phosphosites. An integrated pathway analysis suggested that the increased phosphorylation of CDK7 at Ser164 and Thr170 may serve as the key event in the activation of CDK2 and consequent activation of cell cycle progression. Furthermore, the decrease in GIT1 and WAVE2 protein levels were connected to the disorganization of the actin cytoskeleton, reduction of lamellipodia formation at the leading edge of HaCaT cells, and decrease in their migration capacity. • The Olfactomedin-4 (OLFM4) is present in the basal layer of normal human epidermis. • OLFM4 silencing by siRNA boosts proliferation and inhibits migration of HaCat cells. • Phosphoproteomic analysis unveiled OLFM4 regulated proliferation via CDK7 and CDK2. • The silencing of OLFM4 reduced GIT1 protein level triggering migration inhibition. • OLFM4 is expected to manage keratinocyte migration, proliferation in wound healing. [ABSTRACT FROM AUTHOR]