| [return to list] |
|
Barry
T.
Hinton
Degree(s): Ph.D. Graduate School: The Babraham Institute, Cambridge, UK Primary Appointment: Professor of Cell Biology Research Interests: Cell And Molecular Biology of Epididymal Function Email Address: bth7c@virginia.edu |
|
Biomedical Sciences Graduate Program(s) Research Description
The goal of this laboratory is to understand the mechanisms by which the epididymal
epithelium is able to maintain a specialized luminal fluid microenvironment that
is critically important for the maturation and survival of spermatozoa. This laboratory
utilizes in vivo micropuncture and microperfusion techniques, immunochemical,
microanalytical and modern molecular biological techniques to achieve this goal.
There are four major projects currently ongoing:
(1) To test the hypothesis that growth factors of Sertoli cell origin enter
the epididymal duct and bind to their receptors on the apical surface of the
epididymal cells. Second messengers are then activated which in turn activate
transcription factors that transactivate specific epididymal genes. The gene
of interest in this laboratory is gamma-glutamyl transpeptidase (GGT) which
plays a role in the metabolism of glutathione, an important antioxidant. Multiple
GGTs are expressed in the epididymis and each are under the control of specific
promoters. GGT mRNA IV is of special interest because it is regulated by testicular
factors. Our published and unpublished data has led us to hypothesize that fibroblast
growth factors (FGF) from Sertoli cells interact with specific FGF receptors
(FGFR-1) localized on the apical surface of epididymal cells, activate the MAP
kinase pathway which in turn activate members of Ets transcription factor family,
in particular PEA3, ERM and ER81. These transcription factors then transactivate
GGT mRNA IV. We are using a novel in vivo electroporation technique to examine
the transactivation of GGT IV promoter and laser capture techniques to identify
the FGFR types in specific epididymal cells. We are currently investigating
those proteins, e.g. Grb2 and FRS2, that bind to FGFR-1 and regulate its activity. (2) Fluid homeostasis is critical for normal development and function of the
male reproductive system particularly the epididymis. Alterations in fluid balance
can have adverse effects and may be the source of certain forms of male infertility.
Our laboratory is interested in the expression and regulation of transport proteins
responsible for regulating the epididymal fluid microenvironment. We are currently
studying the expression and regulation of a transport protein that may be responsible
for transporting L-carnitine into the lumen of the epididymis. L-carnitine is
present in high concentrations in the epididymal tissues of many species and
studies suggest that L-carnitine may be involved in water homeostasis. L-carnitine
is transported against a concentration gradient greater than 2000-fold, with
intraluminal concentrations reaching as high as 50mM in the cauda epididymides
of the rat. Studies in our laboratory have demsontrated that OCTN2, a member
of the organic cation transporter family, is expressed in the rat epididymis
in a region-dependent manner with highest expression observed in the regions
previously shown to be involved in L-carnitine uptake. Moreover, we localized
OCTN2 protein to the basolateral region of epididymal cells suggesting that
this protein is likely responsible for the transport of L-carnitine across the
epididymal epithelium. We are currently using gene-silencing technologies to
test the hypothesis that OCTN2 is responsible for transporting L carnitine into
the lumen of the epididymis. Further, we have been performing experiments testing
the hypothesis that OCTN2 and the apical organic solute transporters, FLIPT1
and 2 are regulated by changes in osmolarity. (3) We are investigating the role of the orphan receptor protein tyrosine
kinase, c-Ros, in the post-meiotic, epididymal maturation of sperm. This gene
is involved in the development of the initial segment (IS) of the epididymis.
Male mice with a knock-out mutation in the gene for c-Ros are infertile, and
have IS's that are small and poorly developed; however, these mice appear to
be normal in all other aspects. As expected, c-Ros is expressed in the developing
IS, as well as several other tissues during embryogenesis. However, c Ros may
play a role in the sperm maturation process since this gene is expressed at
high levels in the IS of adult male mice. We are attempting to experimentally
separate these two roles of c-Ros to determine the function of c-Ros in the
adult male. Since the ligand for this receptor is unknown and there are no specific
inhibitors for the protein kinase, we are generating conditional knock-out mice
to regulate the expression of the c-Ros gene in a temporal fashion. We will
allow the IS to develop normally through puberty, and then turn off the c-Ros
gene in the adult. We will then analyze the resulting phenotype. We are also
interested in the signalling pathways and downstream genes controlled by this
receptor protein tyrosine kinase, as well as interacting adaptor proteins and
phosphatases. (4) We have begun experiments to understand the regulation of tubular morphogenesis
of the Wolffian/Mesonephric duct. At E14 the mouse developing Wolffian duct
is approximately 1 mm in length and grows to approximately 1 m. The mechanisms
by which this tube undergoes proliferation and differentiation is unknown. Preliminary
evidence would suggest that growth factors and the hedeghog signaling pathway
are required for normal growth, folding and differentiation. Selected Publications Intranet Profile [To add/update Intranet profile information, read these instructions.]
|
|||||||||||||||