Probes for INS

Background Perturbations to embryonic hemodynamics are known to adversely affect cardiovascular development. Vitelline vein ligation (VVL) is a model of reduced placental blood flow used to induce cardiac defects in early chick embryo development. The effect of these hemodynamic interventions on maturing elastic arteries is largely unknown. We hypothesize that hemodynamic changes impact maturation of the dorsal aorta (DA). Results We examined the effects of VVL on hemodynamic properties well into the maturation process and the corresponding changes in aortic dimensions, wall composition, and gene expression. In chick embryos, we found that DA blood velocity was reduced immediately post-surgery at Hamburger-Hamilton stage (HH) 18 and later at HH36, but not in the interim. Throughout this period, DA diameter adapted to maintain a constant shear stress. At HH36, we found that VVL DAs showed a substantial decrease in elastin and modest increase in collagen protein content. In VVL DAs, upregulation of elastic fiber related genes followed the downregulation of flow-dependent genes. Together, these suggest the existence of a compensatory mechanism in response to shear induced delays in maturation. Conclusions The DA's response to hemodynamic perturbations invokes coupled mechanisms for shear regulation and matrix maturation, potentially impacting the course of vascular development.

In response to cortical stroke and unilateral corticospinal tract degeneration, compensatory sprouting of spared corticospinal fibers is associated with recovery of skilled movement in rodents. To date, little is known about the molecular mechanisms orchestrating this spontaneous rewiring. In this study, we provide insights into the molecular changes in the spinal cord tissue after large ischemic cortical injury in adult female mice, with a focus on factors that might influence the re-innervation process by contralesional corticospinal neurons. We mapped the area of cervical grey matter re-innervation by sprouting contralesional corticospinal axons after unilateral photothrombotic stroke of the motor cortex in mice using anterograde tracing. The mRNA profile of this re-innervation area was analyzed using whole-genome sequencing to identify differentially expressed genes at selected time points during the recovery process. Bioinformatic analysis revealed two phases of processes: Early after stroke (4-7 days post injury), the spinal transcriptome is characterized by inflammatory processes, including phagocytic processes as well as complement cascade activation. Microglia are specifically activated in the denervated corticospinal projection fields in this early phase. In a later phase (28-42 days post injury), biological processes include tissue repair pathways with up-regulated genes related to neurite outgrowth. Thus, the stroke-denervated spinal grey matter, in particular its intermediate laminae, represents a growth-promoting environment for sprouting corticospinal fibers originating from the contralesional motor cortex. This data set provides a solid starting point for future studies addressing key elements of the post-stroke recovery process, with the goal to improve neuroregenerative treatment options for stroke patients.SIGNIFICANCE STATEMENTWe show that the molecular changes in the spinal cord target tissue of the stroke-affected corticospinal tract are mainly defined by two phases: an early inflammatory phase during which microglia are specifically activated in the target area of re-innervating corticospinal motor neurons; and a late phase during which growth-promoting factors are upregulated which can influence the sprouting response, arborization and synapse formation. By defining for the first time the endogenous molecular machinery in the stroke-denervated cervical spinal grey matter with a focus on promotors of axon growth through the growth-inhibitory adult CNS, this study will serve as a basis to address novel neuroregenerative treatment options for chronic stroke patients.

Abstract

OBJECTIVE:

We investigated how pancreatic cancer developed resistance to focal adhesion kinase (FAK) inhibition over time.

DESIGN:

Pancreatic ductal adenocarcinoma (PDAC) tumours from KPC mice (p48-CRE; LSL-KRasG12D/wt; p53flox/wt) treated with FAK inhibitor were analysed for the activation of a compensatory survival pathway in resistant tumours. We identified pathways involved in the regulation of signal transducer and activator of transcription 3 (STAT3) signalling on FAK inhibition by gene set enrichment analysis and verified these outcomes by RNA interference studies. We also tested combinatorial approaches targeting FAK and STAT3 in syngeneic transplantable mouse models of PDAC and KPC mice.

RESULTS:

In KPC mice, the expression levels of phosphorylated STAT3 (pSTAT3) were increased in PDAC cells as they progressed on FAK inhibitor therapy. This progression corresponded to decreased collagen density, lowered numbers of SMA+ fibroblasts and downregulation of the transforming growth factor beta (TGF-β)/SMAD signalling pathway in FAK inhibitor-treated PDAC tumours. Furthermore, TGF-β production by fibroblasts in vitro drives repression of STAT3 signalling and enhanced responsiveness to FAK inhibitor therapy. Knockdown of SMAD3 in pancreatic cancer cells abolished the inhibitory effects of TGF-β on pSTAT3. We further found that tumour-intrinsic STAT3 regulates the durability of the antiproliferative activity of FAK inhibitor, and combinatorial targeting of FAK and Janus kinase/STAT3 act synergistically to suppress pancreatic cancer progression in mouse models.

CONCLUSION:

Stromal depletion by FAK inhibitor therapy leads to eventual treatment resistance through the activation of STAT3 signalling. These data suggest that, similar to tumour-targeted therapies, resistance mechanisms to therapies targeting stromal desmoplasia may be critical to treatment durability.