Incidence and Risk Factors for New and Recurrent Infarcts in Adults With Sickle Cell Disease.

BACKGROUND: Most adults with sickle cell disease will experience a silent cerebral infarction (SCI) or overt stroke. Identifying patient subgroups with increased stroke incidence is important for future clinical trials focused on stroke prevention. Our 3-center prospective cohort study tested the primary hypothesis that adults with sickle cell disease and SCIs have a greater incidence of new stroke or SCI compared with those without SCI. A secondary aim focused on identifying additional risk factors for progressive infarcts, particularly traditional risk factors for stroke in adults. METHODS AND RESULTS: This observational study included adults with sickle cell disease and no history of stroke. Magnetic resonance imaging scans of the brain completed at baseline and >1 year later were reviewed by 3 radiologists for baseline SCIs and new or progressive infarcts on follow-up magnetic resonance imaging. Stroke risk factors were abstracted from the medical chart. Time-to-event analysis was utilized for progressive infarcts. Median age was 24.1 years; 45.3% of 95 participants had SCIs on baseline magnetic resonance imaging. Progressive infarcts were present in 17 participants (17.9%), and the median follow-up was 2.1 years. Incidence of new infarcts was 11.95 per 100 patient-years (6.17-20.88) versus 3.74 per 100 patient-years (1.21-8.73) in those with versus without prior SCI. Multivariable Cox regression showed that baseline SCI predicts progressive infarcts (hazard ratio, 3.46 [95% CI, 1.05-11.39]; P=0.041); baseline hypertension was also associated with progressive infarcts (hazard ratio, 3.23 [95% CI, 1.16-9.51]; P=0.025). CONCLUSIONS: Selecting individuals with SCIs and hypertension for stroke prevention trials in sickle cell disease may enrich the study population with those at highest risk for infarct recurrence.

Loss of flow responsive Tie1 results in ImpairedAortic valve remodeling.

The mechanisms regulating endothelial cell response to hemodynamic forces required for heart valve development, especially valve remodeling, remain elusive. Tie1, an endothelial specific receptor tyrosine kinase, is up-regulated by oscillating shear stress and is required for lymphatic valve development. In this study, we demonstrate that valvular endothelial Tie1 is differentially expressed in a dynamic pattern predicted by disturbed flow during valve remodeling. Following valvular endocardial specific deletion of Tie1 in mice, we observed enlarged aortic valve leaflets, decreased valve stiffness and valvular insufficiency. Valve abnormalities were only detected in late gestation and early postnatal mutant animals and worsened with age. The mutant mice developed perturbed extracellular matrix (ECM) deposition and remodeling characterized by increased glycosaminoglycan and decreased collagen content, as well as increased valve interstitial cell expression of Sox9, a transcription factor essential for normal ECM maturation during heart valve development. This study provides the first evidence that Tie1 is involved in modulation of late valve remodeling and suggests that an important Tie1-Sox9 signaling axis exists through which disturbed flows are converted by endocardial cells to paracrine Sox9 signals to modulate normal matrix remodeling of the aortic valve.

Myocardial differentiation is dependent upon endocardial signaling during early cardiogenesis in vitro.

The endocardium interacts with the myocardium to promote proliferation and morphogenesis during the later stages of heart development. However, the role of the endocardium in early cardiac ontogeny remains under-explored. Given the shared origin, subsequent juxtaposition, and essential cell-cell interactions of endocardial and myocardial cells throughout heart development, we hypothesized that paracrine signaling from the endocardium to the myocardium is crucial for initiating early differentiation of myocardial cells. To test this, we generated an in vitro, endocardial-specific ablation model using the diphtheria toxin receptor under the regulatory elements of the Nfatc1 genomic locus (NFATc1-DTR). Early treatment of NFATc1-DTR mouse embryoid bodies with diphtheria toxin efficiently ablated endocardial cells, which significantly attenuated the percentage of beating EBs in culture and expression of early and late myocardial differentiation markers. The addition of Bmp2 during endocardial ablation partially rescued myocyte differentiation, maturation and function. Therefore, we conclude that early stages of myocardial differentiation rely on endocardial paracrine signaling mediated in part by Bmp2. Our findings provide novel insight into early endocardial-myocardial interactions that can be explored to promote early myocardial development and growth.

What chick and mouse models have taught us about the role of the endocardium in congenital heart disease.

Specific cell and tissue interactions drive the formation and function of the vertebrate cardiovascular system. Although much attention has been focused on the muscular components of the developing heart, the endocardium plays a key role in the formation of a functioning heart. Endocardial cells exhibit heterogeneity that allows them to participate in events such as the formation of the valves, septation of the outflow tract, and trabeculation. Here we review, the contributions of the endocardium to cardiovascular development and outline useful approaches developed in the chick and mouse that have revealed endocardial cell heterogeneity, the signaling molecules that direct endocardial cell behavior, and how these insights have contributed to our understanding of cardiovascular development and disease.

Activation of nuclear factor kappa B in mammary epithelium promotes milk loss during mammary development and infection.

We investigated whether nuclear factor kappa B (NF-kappaB), which exhibits a regulated pattern of activity during murine mammary gland development, plays an important role during lactation and involution, when milk production ceases and the gland undergoes apoptosis and re-modeling. We generated a doxycycline inducible transgenic mouse model to activate NF-kappaB specifically in the mammary epithelium through expression of a constitutively active form of IKK2, the upstream kinase in the classical NF-kappaB signaling cascade. We found that activation of NF-kappaB during involution resulted in a more rapid reduction in milk levels and increased cleavage of caspase-3, an indicator of apoptosis. We also found that activation of NF-kappaB during lactation with no additional involution signals had a similar effect. The observation that NF-kappaB is a key regulator of milk production led us to investigate the role of NF-kappaB during mastitis, an infection of the mammary gland in which milk loss is observed. Mammary gland injection of E. coli LPS resulted in activation of NF-kappaB and milk loss during lactation. This milk loss was decreased by selective inhibition of NF-kappaB in mammary epithelium. Together, our data reveal that activation of NF-kappaB leads to milk clearance in the lactating mammary gland. Therefore, targeting of NF-kappaB signaling may prove therapeutic during mastitis in humans and could be beneficial for the dairy industry, where such infections have a major economic impact.

The nuclear factor-kappaB pathway controls the progression of prostate cancer to androgen-independent growth.

Typically, the initial response of a prostate cancer patient to androgen ablation therapy is regression of the disease. However, the tumor will progress to an "androgen-independent" stage that results in renewed growth and spread of the cancer. Both nuclear factor-kappaB (NF-kappaB) expression and neuroendocrine differentiation predict poor prognosis, but their precise contribution to prostate cancer progression is unknown. This report shows that secretory proteins from neuroendocrine cells will activate the NF-kappaB pathway in LNCaP cells, resulting in increased levels of active androgen receptor (AR). By blocking NF-kappaB signaling in vitro, AR activation is inhibited. In addition, the continuous activation of NF-kappaB signaling in vivo by the absence of the IkappaBalpha inhibitor prevents regression of the prostate after castration by sustaining high levels of nuclear AR and maintaining differentiated function and continued proliferation of the epithelium. Furthermore, the NF-kappaB pathway was activated in the ARR(2)PB-myc-PAI (Hi-myc) mouse prostate by cross-breeding into a IkappaBalpha(+/-) haploid insufficient line. After castration, the mouse prostate cancer continued to proliferate. These results indicate that activation of NF-kappaB is sufficient to maintain androgen-independent growth of prostate and prostate cancer by regulating AR action. Thus, the NF-kappaB pathway may be a potential target for therapy against androgen-independent prostate cancer.

A transgenic model reveals important roles for the NF-kappa B alternative pathway (p100/p52) in mammary development and links to tumorigenesis.

A regulated pattern of nuclear factor kappaB (NF-kappaB) activation is essential for normal development of the mammary gland. An increase in NF-kappaB activity has been implicated in breast cancer. We have generated a novel transgenic mouse model to investigate the role of the alternative NF-kappaB pathway in ductal development and identify possible mediators of tumorigenesis downstream of p100/p52. By overexpressing the NF-kappaB p100/p52 subunit in mammary epithelium using the beta-lactoglobulin milk protein promoter, we found that transgene expression resulted in increased overall NF-kappaB activity during late pregnancy. During pregnancy, p100/p52 expression resulted in delayed ductal development with impaired secondary branching and increased levels of Cyclin D1, matrix metalloproteinase-2 (MMP-2), matrix metalloproteinase-9 (MMP-9), and cyclo-oxygenase-2 (COX-2) in the mammary gland. After multiple pregnancies the p100 transgenics exhibited a ductal thickening accompanied by small hyperplastic foci. In tumors from mice expressing the polyoma middle T oncoprotein (PyVT) in the mammary gland, increased levels of p100/p52 were present at the time of tumor development. These results show that increased p100/p52 disrupts normal ductal development and provides insight into the mechanism by which this may contribute to human breast cancer.