Keratin 19 interacts with GSK3β to regulate its nuclear accumulation and degradation of cyclin D3

Keratin 19 (K19) inhibits glycogen synthase kinase-3β (GSK3β) accumulation in the nucleus, preventing cyclin D3 degradation. K19 physically interacts with GSK3β, and this interaction requires Ser 10 and 35 of K19. Mutating either residues decreased cyclin D3 levels and cell proliferation. The K19–GSK3β–cyclin D3 pathway also regulates the sensitivity of cancer cells to CDK4/6 inhibitors.

Keratin 19 interacts with GSK3β to regulate its nuclear accumulation and degradation of cyclin D3

Cyclin D3 regulates the G1/S transition and is frequently overexpressed in several cancer types including breast cancer, where it promotes tumor progression. Here, we show that a cytoskeletal protein keratin 19 (K19) physically interacts with a serine/threonine kinase GSK3β and prevents GSK3β-dependent degradation of cyclin D3. The absence of K19 allowed active GSK3β to accumulate in the nucleus and degrade cyclin D3. Specifically, the head domain of K19 was required to sustain inhibitory phosphorylation of GSK3β Ser9, prevent nuclear accumulation of GSK3β, and maintain cyclin D3 levels and cell proliferation. K19 was found to interact with GSK3β and K19-GSK3β interaction was mapped out to require Ser10 and Ser35 residues on the head domain of K19. Unlike wildtype K19, S10A and S35A mutants failed to maintain total and nuclear cyclin D3 levels and induce cell proliferation. Finally, we show that the K19-GSK3β-cyclin D3 pathway affected sensitivity of cells towards inhibitors to cyclin dependent kinase 4 and 6 (CDK4/6). Overall, these findings establish a role for K19 in the regulation of GSK3β-cyclin D3 pathway and demonstrate a potential strategy for overcoming resistance to CDK4/6 inhibitors.

The potential of mesenchymal stem‐cell secretome for regeneration of intervertebral disc: A review article

Low back pain is a crucial public health problem that is commonly associated with intervertebral disc de‐ generation and has vast socio‐economic impact worldwide. Current treatments for disc degeneration are conservative, non‐surgical, or surgical interventions, and there is no current clinical therapy aimed at directly reversing the degeneration. Given the limited capacity of intervertebral disc (IVD) cells to self‐repair, treatment aiming to regenerate IVDs is a topic of interest and mesenchymal stem cells (MSCs) have been identified as having potential in this regeneration. Recent studies have revealed that the benefits of MSC therapy could result from the molecules the cells secrete and that play principal roles in regulating essential biologic processes, rather than from the implanted cells themselves. Therefore, the objective of this study is to review the potential use of the MSC secretome to regenerate IVDs. Current evidence shows that the secretome may regenerate IVDs by modulating the gene expressions of nucleus pulposus cells (upregulation of keratin 19 and downregulation of matrix metalloproteinase 12 and matrix Gla protein) and stimulating IVD progenitor cells to repair the degenerated disc.

Mesothelial cells are not a source of adipocytes in mice

Visceral adipose tissue (VAT) depots are associated with the adverse metabolic consequences of obesity, such as insulin resistance. The developmental origin of VAT depots and the identity and regulation of adipocyte progenitor cells have been active areas of investigation. In recent years, a paradigm of mesothelial cells as a source of VAT adipocyte progenitor cells has emerged based on lineage-tracing studies using the Wilms' tumor gene, Wt1, as a marker for cells of mesothelial origin. Here we show that Wt1 expression in adipose tissue is not limited to the mesothelium, but is also expressed by a distinct preadipocyte population in both mice and humans. We identify keratin 19 (Krt19) as a highly-specific marker for the adult mouse mesothelium, and demonstrate that Krt19-expressing mesothelial cells do not differentiate into visceral adipocytes. These results contradict the assertion that the VAT mesothelium can serve as a source of adipocytes.

Mesenteric and Retroperitoneal Mucinous Cystic Neoplasms: A Case Series

Aims. Mucinous cystic neoplasms (MCNs) are cystic neoplasms with mucinous epithelium surrounded by ovarian-like stroma. Extraovarian MCN occurring in the liver and pancreas have been well characterized. However, only rare case reports of MCN arising outside of these locations have been reported. MCNs arising in unusual locations should enter the differential diagnosis of mucinous intra-abdominal tumors and must be distinguished from more common mimics. Therefore, we aimed to examine a series of MCNs of the retroperitoneum and mesentery to characterize the clinicopathologic features of this entity. Methods and results. Seven MCNs arising in the abdominal mesentery or retroperitoneum were retrospectively identified. A clinicopathologic, histologic, and immunohistochemical (keratin 7, keratin 19, keratin 20, calretinin, inhibin-α, steroidogenic factor-1 (SF-1), estrogen receptor (ER), progesterone receptor (PR), PAX8, CDX2, and CD10) analysis was performed. All 7 MCNs were from females with a median age of 41 years old and a median size of 8 cm. All cases demonstrated mucinous with or without concomitant non-mucinous epithelium overlying spindle cell ovarian-like stroma. Luteinized cells were noted. The epithelium was positive for keratin 7 and keratin 19 in all 7 cases, while the stroma expressed ER, PR, and SF-1 in all cases stained. Calretinin was focally positive in the stroma of 3 of 7 cases, while inhibin-α was focally expressed in 5 of 6 cases. Conclusions. These results highlight the clinicopathologic, histologic, and immunophenotypic similarities between MCNs of the mesentery, retroperitoneum, pancreas, and liver. Overlapping features suggest a common histogenesis for all MCNs, which could include periductal fetal mesenchyme, aberrant migration of primordial germ cells, or abnormal differentiation or metaplasia of the embryonic coelomic epithelium.

Non-viral reprogramming of human nucleus pulposus cells with FOXF1 via extracellular vesicle delivery: an in vitro and in vivo study

Intervertebral disc (IVD) degeneration is characterized by decreased cellularity and proteoglycan synthesis and increased inflammation, catabolism, and neural/vascular ingrowth. Regenerative methods for IVD degeneration are largely cell-therapy-based or involve viral vectors, which are associated with mutagenesis and undesired immune responses. The present study used bulk electroporation and engineered extracellular vesicles (EVs) to deliver forkhead-box F1 (FOXF1) mRNA to degenerate human nucleus pulposus (NP) cells as a minimally invasive therapeutic strategy for IVD regeneration. Bulk electroporation was used to investigate FOXF1 effects on human NP cells during a 4-week culture in 3D agarose constructs. Engineered EV delivery of FOXF1 into human IVD cells in monolayer was determined, with subsequent in vivo validation in a pilot mouse IVD puncture model. FOXF1 transfection significantly altered gene expression by upregulating healthy NP markers [FOXF1, keratin 19 (KRT19)], decreasing inflammatory cytokines [interleukin (IL)-1β, -6], catabolic enzymes [metalloproteinase 13 (MMP13)] and nerve growth factor (NGF), with significant increases in glycosaminoglycan accumulation in human NP cells. Engineered EVs loaded with FOXF1 demonstrated successful encapsulation of FOXF1 cargo and effective uptake by human NP cells cultured in monolayer. Injection of FOXF1-loaded EVs into the mouse IVD in vivo resulted in a significant upregulation of FOXF1 and Brachyury, compared to controls at 7 d post-injection, with no evidence of cytotoxicity. This is the first study to demonstrate non-viral delivery of FOXF1 and reprogramming of human NP cells in vitro and mouse IVD cells in vivo. This strategy represents a non-addictive approach for treating IVD degeneration and associated back pain.