The Roles of Bone Marrow-Derived Stem Cells in Coronary Collateral Growth Induced by Repetitive Ischemia.

Many clinical trials have attempted to use stem cells to treat ischemic heart diseases (IHD), but the benefits have been modest. Though coronary collaterals can be a "natural bypass" for IHD patients, the regulation of coronary collateral growth (CCG) and the role of endogenous stem cells in CCG are not fully understood. In this study, we used a bone marrow transplantation scheme to study the role of bone marrow stem cells (BMSCs) in a rat model of CCG. Transgenic GFP rats were used to trace BMSCs after transplantation; GFP bone marrow was harvested or sorted for bone marrow transplantation. After recovering from transplantation, the recipient rats underwent 10 days of repetitive ischemia (RI), with echocardiography before and after RI, to measure cardiac function and myocardial blood flow. At the end of RI, the rats were sacrificed for the collection of bone marrow for flow cytometry or heart tissue for imaging analysis. Our study shows that upon RI stimulation, BMSCs homed to the recipient rat hearts' collateral-dependent zone (CZ), proliferated, differentiated into endothelial cells, and engrafted in the vascular wall for collateral growth. These RI-induced collaterals improved coronary blood flow and cardiac function in the recipients' hearts during ischemia. Depletion of donor CD34+ BMSCs led to impaired CCG in the recipient rats, indicating that this cell population is essential to the process. Overall, these results show that BMSCs contribute to CCG and suggest that regulation of the function of BMSCs to promote CCG might be a potential therapeutic approach for IHD.

Comparative embryology of Delphinapterus leucas (beluga whale), Balaena mysticetus (bowhead whale), and Stenella attenuata (pan-tropical spotted dolphin) (Cetacea: Mammalia).

Embryogenesis of cetaceans (whales, dolphins, porpoises) is best known in Stenella attenuata, the pan-tropical spotted dolphin, based on a remarkably complete and well-studied prenatal ontogenetic series. Our study expands understanding of cetacean embryology by adding two additional cetacean taxa: the beluga whale (Delphinapterus leucas, Odontoceti), and the bowhead whale (Balaena mysticetus, Mysticeti). We identify key features that characterize these taxa at specific stages and highlight heterochrony between the odontocetes and mysticetes. The toothed whales are more similar in developmental timing to each other than either is to Balaena. The two odontocete taxa, Stenella and Delphinapterus, share similar developmental trajectories while early Balaena specimens differ from the odontocetes. This developmental variation proves challenging to ascribe to the existing Carnegie staging system. Most notably, flippers, hindlimbs, and flukes all provide morphological traits for characterization within the Carnegie staging system. A presomitic Delphinapterus embryo is also described. This study applies the Carnegie staging system to two more cetacean taxa and forms a framework for future research on cetacean developmental genetics and the modeling of fetal growth.

Aquatic Habits of Cetacean Ancestors: Integrating Bone Microanatomy and Stable Isotopes.

The earliest cetaceans were interpreted as semi-aquatic based on the presence of thickened bones and stable oxygen isotopes in tooth enamel. However, the origin of aquatic behaviors in cetacean relatives (e.g., raoellids, anthracotheres) remains unclear. This study reconstructs the origins of aquatic behaviors based on long bone microanatomy and stable oxygen isotopes of tooth enamel in modern and extinct cetartiodactylans. Our findings are congruent with published accounts that microanatomy can be a reliable indicator of aquatic behaviors in taxa that are obligatorily aquatic, and also highlight that some "semi-aquatic" behaviors (fleeing into the water to escape predation) may have a stronger relationship to bone microanatomy than others (herbivory in near-shore aquatic settings). Bone microanatomy is best considered with other lines of information in the land-to-sea transition of cetaceans, such as stable isotopes. This study extends our understanding of the progression of skeletal phenotypes associated with habitat shifts in the relatives of cetaceans.

Severe Bone Loss as Part of the Life History Strategy of Bowhead Whales.

The evolution of baleen constituted a major evolutionary change that made it possible for baleen whales to reach enormous body sizes while filter feeding on tiny organisms and migrating over tremendous distances. Bowhead whales (Balaena mysticetus) live in the Arctic where the annual cycle of increasing and decreasing ice cover affects their habitat, prey, and migration. During the nursing period, bowheads grow rapidly; but between weaning and approximately year 5, bowhead whales display sustained baleen and head growth while limiting growth in the rest of their bodies. During this period, they withdraw resources from the skeleton, in particular the ribs, which may lose 40% of bone mass. Such dramatic changes in bones of immature mammals are rare, although fossil cetaceans between 40 and 50 million years ago show an array of rib specializations that include bone loss and are usually interpreted as related to buoyancy control.

Organization and distribution of glomeruli in the bowhead whale olfactory bulb.

Although modern baleen whales (Mysticeti) retain a functional olfactory system that includes olfactory bulbs, cranial nerve I and olfactory receptor genes, their olfactory capabilities have been reduced to a great degree. This reduction likely occurred as a selective response to their fully aquatic lifestyle. The glomeruli that occur in the olfactory bulb can be divided into two non-overlapping domains, a dorsal domain and a ventral domain. Recent molecular studies revealed that all modern whales have lost olfactory receptor genes and marker genes that are specific to the dorsal domain. Here we show that olfactory bulbs of bowhead whales (Balaena mysticetus) lack glomeruli on the dorsal side, consistent with the molecular data. In addition, we estimate that there are more than 4,000 glomeruli elsewhere in the bowhead whale olfactory bulb, which is surprising given that bowhead whales possess only 80 intact olfactory receptor genes. Olfactory sensory neurons that express the same olfactory receptors in rodents generally project to two specific glomeruli in an olfactory bulb, implying an approximate 1:2 ratio of the number of olfactory receptors to the number of glomeruli. Here we show that this ratio does not apply to bowhead whales, reiterating the conceptual limits of using rodents as model organisms for understanding the initial coding of odor information among mammals.

Seasonal and Ontogenetic Variation in Subcutaneous Adipose Of the Bowhead Whale (Balaena mysticetus).

Cetacean evolution was shaped by an extraordinary land-to-sea transition in which the ancestors of whales became fully aquatic. As part of this transition, these mammals evolved unusually thick blubber which acts as a metabolic reservoir as well as an insulator and provides buoyancy and streamlining. This study describes blubber stratification and correlates it to seasonal variation, feeding patterns, and ontogeny in an arctic-adapted mysticete, the bowhead whale (Balaena mysticetus). Bowheads are unique among mammals for possessing the largest known blubber stores. We found that adipocyte numbers in bowheads, like other mammals, do not vary with season or feeding pattern but that adipocyte size and structural fiber densities do vary with blubber depth.

The spiral ganglion and Rosenthal's canal in beluga whales.

With the increase of human activity and corresponding increase in anthropogenic sounds in marine waters of the Arctic, it is necessary to understand its effect on the hearing of marine wildlife. We have conducted a baseline study on the spiral ganglion and Rosenthal's canal of the cochlea in beluga whales (Delphinapterus leucas) as an initial assessment of auditory anatomy and health. We present morphometric data on the length of the cochlea, number of whorls, neuron densities along its length, Rosenthal's canal length, and cross-sectional area, and show some histological results. In belugas, Rosenthal's canal is not a cylinder of equal cross-sectional area, but its cross-section is greatest near the apex of the basal whorl. We found systematic variation in the numbers of neurons along the length of the spiral ganglion, indicating that neurons are not dispersed evenly in Rosenthal's canal. These results provide data on functionally important structural parameters of the beluga ear. We observed no signs of acoustic trauma in our sample of beluga whales.

Delineation of VEGF-regulated genes and functions in the cervix of pregnant rodents by DNA microarray analysis.

BACKGROUND: VEGF-regulated genes in the cervices of pregnant and non-pregnant rodents (rats and mice) were delineated by DNA microarray and Real Time PCR, after locally altering levels of or action of VEGF using VEGF agents, namely siRNA, VEGF receptor antagonist and mouse VEGF recombinant protein. METHODS: Tissues were analyzed by genome-wide DNA microarray analysis, Real-time and gel-based PCR, and SEM, to decipher VEGF function during cervical remodeling. Data were analyzed by EASE score (microarray) and ANOVA (Real Time PCR) followed by Scheffe's F-test for multiple comparisons. RESULTS: Of the 30,000 genes analyzed, about 4,200 genes were altered in expression by VEGF, i.e., expression of about 2,400 and 1,700 genes were down- and up-regulated, respectively. Based on EASE score, i.e., grouping of genes according to their biological process, cell component and molecular functions, a number of vascular- and non-vascular-related processes were found to be regulated by VEGF in the cervix, including immune response (including inflammatory), cell proliferation, protein kinase activity, and cell adhesion molecule activity. Of interest, mRNA levels of a select group of genes, known to or with potential to influence cervical remodeling were altered. For example, real time PCR analysis showed that levels of VCAM-1, a key molecule in leukocyte recruitment, endothelial adhesion, and subsequent trans-endothelial migration, were elevated about 10 folds by VEGF. Further, VEGF agents also altered mRNA levels of decorin, which is involved in cervical collagen fibrillogenesis, and expression of eNO, PLC and PKC mRNA, critical downstream mediators of VEGF. Of note, we show that VEGF may regulate cervical epithelial proliferation, as revealed by SEM. CONCLUSION: These data are important in that they shed new insights in VEGF's possible roles and mechanisms in cervical events near-term, including cervical remodeling.

Glutamate and metabotropic glutamate receptors associated with innervation of the uterine cervix during pregnancy: receptor antagonism inhibits c-Fos expression in rat lumbosacral spinal cord at parturition.

Dorsal root ganglia (DRG) neurons connect the spinal cord and uterine cervix, and are activated at parturition with subsequent stimulation of secondary neurons in the spinal dorsal horn and autonomic areas. Neuropeptide neurotransmitters and receptors have been studied in these areas, but amino acid transmitters, e.g., glutamate, an excitatory neurotransmitter involved in sensory and nociceptive processing, have not been characterized. To determine if glutamate is involved in innervation of the cervix, rats were examined for markers of glutamatergic neurons in the L6-S1 spinal cord, DRG and cervix. Metabotropic glutamate receptors mGluR5 in the spinal dorsal horn and their expression over pregnancy were examined in pregnant rats and pregnant rats treated continuously with an antagonist of mGluR5, 2-methyl-6-(phenylethynyl) pyridine (MPEP). Rats were allowed to deliver pups to determine if the antagonist altered the expression of an early response gene protein, Fos, in the L6-S1 cord. Immunohistochemistry showed glutamate- and vesicular glutamate transporter1 (VGluT1)-positive fibers in the cervix, glutamate- and VGluT1-expressing neurons in the DRG, some of which also exhibited retrograde tracer from cervical injections, and VGluT1 and mGluR5 immunoreactivities in the L6-S1 spinal dorsal horns. Expression of mGluR5 receptors increased over pregnancy. Fos-positive neurons were present among mGluR5-immunoreactivity in the spinal dorsal horn. Parturition-induced Fos-positive neurons in the spinal cords were abundant in control rats, but were reduced by 70% in MPEP-treated animals. These results suggest that glutamate is likely involved in the transmission of sensory signals, possibly pain, from the cervix to the spinal cord at parturition.