Email: c.warner@neu.edu

Phone: 617.373.4036
Fax: 617.373.3724

Location: 306 E Mugar Life Sciences
Mail: NU/Biology
         134 Mugar Life Sciences
         360 Huntington Avenue
         Boston, MA 02115 USA

Warner Lab

Research Description

The overall aim of my laboratory is to understand the parameters that define the health of preimplantation embryos and embryonic stem cells by using genetic, immunological, and imaging methods.  The preimplantation period of development starts at the moment of fertilization and continues until implantation of the embryo into the uterine wall, a process that takes 5 days in the mouse and 6 days in humans.  During the preimplantation period the embryos are free floating in the reproductive tract. This makes it possible to remove the embryos from their mothers and subject them to well-controlled experimental protocols.  We use the mouse as a model system with the long-range goal of using results from our experiments on mice to develop methods to identify healthy human embryos created in in vitro fertilization (IVF) clinics. Our second long-range goal is to identify which preimplantation embryos have the greatest potential to give rise to healthy embryonic stem cells.  Images of mouse and human preimplantation embryos are shown in Figure 1 and Figure 2, respectively.

Figure 1. Mouse preimplantation embryos.

Figure 2. Human preimplantation embryos.

Basic research on mice in the 1950’s and 1960’s led to the techniques that were used to accomplish IVF in humans. Since the birth of the first baby created by IVF, Louise Brown in England in 1978, a multi-billion dollar IVF industry has emerged that has produced almost two million IVF babies worldwide. In the United States alone there are more than 400 IVF clinics that produce about 75,000 IVF babies each year. However, the success rate of producing live babies after IVF is only about 25%, in large part due to the inability to properly assess embryo quality by present biochemical, genetic, and imaging methods. To increase the rate of pregnancy success most clinics transfer three embryos back to the mothers. This leads to a 20-fold increase in the risk of twins and a 400-fold increased risk of triplets. The downside of multiple births is the very large increase in perinatal mortality and morbidity. For instance, 24% of IVF triplets have cerebral palsy. Moreover, many women undergoing IVF are in their late 30’s or early 40’s, so there is great physical and emotional stress on the mothers as well as increased morbidity and mortality in the babies. Our research is aimed at developing genetic, immunological, and imaging methods to assess preimplantation embryo health so that only one embryo can be transferred back to the mother.

1. Genes that influence preimplantation embryo health.

We have discovered a gene, the Ped gene, which has a major influence on the rate of cleavage of preimplantation mouse embryos and their subsequent chance of survival. We have identified the product of the Ped gene as a protein called Qa-2. Qa-2 is a very interesting molecule because it is part of a family of proteins, the major histocompatibility complex (MHC) class I proteins, which play a vital role in the regulation of the immune response. The finding of a function for a MHC class I protein in early development is novel and exciting. We have found that preimplantation mouse embryos that express Qa-2 protein on their cell surface cleave at a fast rate whereas embryos that are missing Qa-2 protein (due to a gene deletion) cleave at a slow rate. A diagram depicting these two types of embryos is shown in Figure 3.

Figure 3. Mouse embryos with the Ped slow and Ped fast alleles have the absence of presence of Qa-2 on the embryonic surface.

We are applying a number of different approaches to identify the mechanisms by which Qa-2 protein on the cell surface signals embryos to cleave at a fast rate. These include studies to identify partner proteins for Qa-2, studies to identify genes other than the Ped gene that regulate the rate of cleavage, and studies of the expression of the protein products of these genes using various techniques.

In addition, we are conducting experiments aimed at the identification of the human homolog of mouse Qa-2, which is a protein called HLA-G. This is important because it has been established that human embryos that cleave at a fast rate have a higher chance of survival than embryos that cleave at a slow rate after IVF.

2. Genes that influence oocyte death.

If a mouse or human egg (oocyte) is not fertilized it dies. There is recent evidence that the mechanism of this death is by apoptosis. This is an important concept because only death by apoptosis, and not death by necrosis, prevents the release of inflammatory cytokines, which would be deleterious to a subsequent pregnancy. We are determining which genes and proteins are involved in the induction of apoptosis in unfertilized oocytes. An example of the expression of caspase-3, one of the effectors of apoptosis, in normal mouse oocytes and mouse oocytes dying by apoptosis is shown in Figure 4. In the healthy oocyte only the second polar body is stained for caspase-3 activity. In the unhealthy oocyte all of the cell fragments (fragments are indicative of apoptosis) are stained, indicating that the oocyte has died by apoptosis, not necrosis. We are investigating the details of the biochemical pathways that lead to the apoptotic death of unfertilized oocytes, with especial emphasis on the role of mitochondria in oocyte health.

Figure 4. Staining of healthy and unhealthy oocytes for capsase-3, an effector of apoptosis.

3. Imaging to assess oocyte and embryo health.

Which egg will produce a healthy baby? We have built a state-of-the-art non-invasive imaging instrument to assess oocyte and embryo health. This instrument, called the Keck three-dimensional fusion microscope (3DFM), combines five imaging modalities into a single platform so that specimens can be evaluated at the same time and at the same place with five complementary imaging methods. The five modalities are differential interference contrast (DIC) microscopy, laser scanning confocal microscopy (LSCM), reflectance confocal microscopy (RCM), two-photon laser scanning microscopy (TPLSM), and quadrature tomographic microscopy (QTM). The last technique uses an instrument, the QTM, which has been developed by engineers at Northeastern University, and is the only microscope of its kind in the world. A schematic of the Keck 3DFM is shown in Figure 5.

Figure 5. The Keck 3DFM combines microscopes that can be sequentially directed onto a fixed specimen by switching optics without moving the specimen.

As illustrated in Figure 6, we have a major research effort underway to use imaging to distinguish healthy from unhealthy oocytes and embryos.

Figure 6. Which egg will produce a healthy baby?

4.  The Embryo-Stem Cell Loop

Preimplantation embryos have the potential to implant in a uterus leading to the birth of a baby.  At the final stage before implantation, blastocyst stage embryos form two distinct tissue types: the trophectoderm (TE) and the inner cell mass (ICM) (see Figures 1 and 2).  The TE forms the placenta, and the ICM forms the embryo proper.  Thus, the cells in the ICM have the capacity to form any tissue type in the body.  By harvesting these ICM cells, we can establish cultured cell lines of embryonic stem (ES) cells.  ES cells have the potential to form tissue types such as neurons, blood, skin, cardiac muscle, bone, and even germ cells (eggs and sperm) as shown in Figure 7.  There is hope that tissues grown from ES cells may be used therapeutically to alleviate the effects of many types of disorders and diseases such as spinal cord injuries, diabetes, and Parkinson’s disease.  We are very interested in the processes involved in maintaining ES cells in their undifferentiated state and in the mechanisms involved in their differentiation.  We are making use of the Keck 3DFM to study these mechanisms.

Selected Publications

Research in my laboratory is a team effort.  Senior research scientists, postdoctoral fellows, graduate students and undergraduate students participate in the publication of these research results.

Warner, C.M. (2006) Immunological aspects of embryo development. In: J. Cohen and K. Elder (eds.) Human Embryo Evaluation & Selection, Parthenon Publishing Group, United Kingdom, in press

Byrne, M.J., Newmark, J.A., Warner, C.M. (2006) Analysis of the sex ration in preimplantation embryos from B6.K1 and BB6.K2 Ped gene congenic mice. J. Asst. Reprod. Gen., epub Aug 11. [PDF]

Warger II, W.C., Newmark, J.A., Warner, C.M., and DiMarzio, C.A. (2006) Phase subtraction cell counting method for live mouse embryos. Optical Society of America Proceedings, in press.

Purnell, E.T., Warner, C.M., Kort, H.I., Mitchell-Leef, D., Elsner, C.W., Shapiro, D.B., Massey, J.B., and Roudebush, W.E. (2006) Influence of the preimplanntation embryo development (Ped) gene on embryonic platelet-activating factor (PAF) levels. J. Asst. Reprod. Gen., in press.

Warger II, W.C., Newmark, J.A., Zhao, B., Warner, C.M., and DiMarzio, C. A. (2006) Accurate cell counts in live mouse embryos using optical quadrature differential interference contrast microscopy. Proc. SPIE, 6090, 30-41. [PDF]

Watkins, A.J., Wilkins, A., Osmond, C., Warner, C.M., Comiskey, M., Hanson, M., and Fleming, T.P. (2006) The influence of mouse Ped gene expression on postnatal development.  J. Physiol., 571, 211-220. [PDF]

Comiskey, M., Warner, C.M., and Schust, D.J. (2006) MHC molecules of the preimplantation embryo and trophoblast.  In:   G. Mor (ed.) Immunology of Pregnancy, Landes Bioscience, Georgetown, TX, pp. 130-147.

Assounga, A.G. and Warner, C.M. (2005) Memory T lymphocytes of young and old C57BL/6 mice express high levels of class I major histocompatibility complex (H-2 Kb) protein.  Growth, Develop., Aging, 69, 59-66. [PDF]

Warner, C.M., Newmark, J.A., Comiskey, M, De Fazio, S.R., O’Malley, D.M., Rajadhyaksha, M., Townsend, D.J., McKnight S., Roysam, B., Dwyer, P.J., and DiMarzio, C.A.  (2004b)  Genetics and imaging to assess oocyte and preimplantation embryo health.  Reprod., Fertil., Develop. 16, 729-741. [PDF]

Warner, C.M., Comiskey, M., Clisham, P.R., and Brenner, C.A.  (2004)  Soluble HLA-G (sHLA-G)-A predictor of IVF outcome?  J. Asst. Reprod. Gen., 21, 315-316. [PDF]

Assounga, A. and Warner, C.M. (2004)  Transcription of major histocompatibility complex class I (Kb) associated with antigen processing 1 and 2 genes is up-regulated with age.  Immunology, 113, 378-83. [PDF]

Papandile, A., Tyas, D., O'Malley, D.M., and Warner, C.M. (2004) Analysis of Capase-3, capase-8 and capase-9 enzymatic activities in mouse oocytes and zygotes. Zygote, 12, 57-64. [PDF]

Townsend, D.J., Quarles, K.D., Thomas, A.L., Rockward, W.S., Warner, C.M., Newmark, J.A., and DiMarzio, C.A. (2003) Quantitative phase measurements using a Quadrature Tomographic Microscope. SPIE Proceedings, 59-65. [PDF]

Wu, H., Norum, B., Newmark, J., Salzberg, B., Warner, C.M., DiMarzio, C., and Kaeli, D. (2003) The CenSSIS image database. Proceedings of the 15th International Conference on Scientific and Statistical Databases, IEEE Press, pp. 117-126. [PDF]

Comiskey, M., Goldstein, C.Y., De Fazio, S.R., Mammolenti, M., Newmark, J.A., and Warner, C.M. (2003) Evidence that HLA-G is the functional homolog of Qa-2, the Ped gene product. Hum. Immunol., 64, 999-1004. [PDF]

Newmark, J.A., Sacher, F., Jones, G.S., and Warner, C.M. (2002) Ped gene deletion polymorphism frequency in wild mice. J. Exptl. Zool., 293, 179-185.

Gordo, A.C., Rodrigues, P., Kurokawa, M., Jellerette, T., Exley, G.E., Warner, C.M., and Fissore, R. (2002) Intracellular calcium oscillations signal apoptosis rather than activation in in vitro aged mouse eggs. Biol. Reprod., 66, 1828-1837. [PDF]

Warner, C.M., Tyas, D.A., Goldstein, C.S., Comiskey, M., Cohen, J., and Brenner, C.A. (2002) Genotyping: The HLA system and embryo development. Reproductive BioMedicine Online, 4, 133-139. [PDF]

Warner, C.M. and Brenner, C.A. (2001) Genetic regulation of preimplantation embryo survival. Current Topics in Developmental Biology, 52, 151-192.

Stott, J., Bennett, E., Warner, C.M., and DiMarzio, C.A. (2001) Three-dimensional imaging with a Quadrature Tomographic Microscope. SPIE Proceedings, 4261, 24-32. [PDF]

McElhinny, A.S. and Warner, C.M. (2000) Crosslinking of Qa-2 protein, the Ped gene product, increases the cleavage rate of C57BL/6 mouse embryos. Mol. Human Reprod., 6, 517-522. [PDF]

McElhinny, A.S., Exley, G.E., and Warner, C.M. (2000) Painting Qa-2 onto Ped slow preimplantation embryos increases their rate of cleavage. Amer. J. Reprod. Immunol., 44, 52-58. [PDF]

Warner, C.M. and Paschetto, M. G. (2000) Analysis of mRNA levels for the MHC classI-like molecules CD1 and FcRn in preimplantation mouse embryos. Amer. J. Reprod. Immunol., 43, 234-239. [PDF]

Ke, X. and Warner, C.M. (2000) Regulation of Ped gene expression by TAP protein. J. Reprod. Immunol., 46, 1-15. [PDF]

Glina, Y., Tsihrintzes, G.A., Warner, C.M., Hogenboom, D.O., and DiMarzio, C.A. (1999) On the use of the optical quadrature method in tomographic microscopy. SPIE Proceedings, 3605, 101-106. [PDF]

Cohen, J., Brenner, C.A., Warner, C., Steuerwald, N., Sadowy, S., Barritt, J., Sandalinas, M., and Munne, S. (1999) Genetics of the fertilizing egg. In “Towards Reproductive Certainty: Fertility and Genetics Beyond 1999.” Jansen, R. and Mortimer, D. (eds.), pp. 231-246, Parthenon Publishing Group, New York. [PDF]

Cao, W., Brenner, C.A., Alikani, M., Cohen, J., and Warner, C.M. (1999) Search for a human homolog of the mouse Ped gene. Mol. Human Reprod., 5, 541-547. [PDF]

Exley, G.E., Tang, C., McElhinny, A.S., and Warner, C.M. (1999) Expression of caspase and BCL-2 apoptotic family members in mouse preimplantation embryos. Biol. Reprod., 61, 231-239. [PDF]

Wu, L., Feng, H., and Warner, C.M. (1999) Identification of two major histocompatibility complex class Ib genes, Q7 and Q9, as the Ped gene in the mouse. Biol. Reprod., 60, 1114-1119. [PDF]

Exley, G.E. and Warner, C.M. (1999b) Selection in favor of the Ped fast haplotype occurs between mid-gestation and birth. Immunogenetics, 49, 653-659. [PDF]

Wu, L., Exley, G.E., and Warner, C.M. (1998) Differential expression of Ped gene candidates in preimplantation mouse embryos. Biol. Reprod., 59, 941-952. [PDF]

Warner, C.M., Exley, G.E., McElhinny, A.S., and Tang, C. (1998c) Genetic regulation of preimplantation mouse embryo survival. J. Exptl. Zool., 282, 272-279. [PDF]

Warner, C.M., Cao, W., Exley, G.E., McElhinny, A.S., Alikani, M., Cohen, J., Scott, R.T., and Brenner, C.A. (1998b) Genetic regulation of egg and embryo survival. Human Reprod., 13, 178-190. [PDF]

Warner, C.M., McElhinny, A.S., Wu, L., Cieluch, C., Ke, X., Cao, W., Tang, C., and Exley, G.E. (1998) Role of the Ped gene and apoptosis genes in control of preimplantation development. J. Asst. Reprod. Gen., 331-337. [PDF]

McElhinny, A.S., Kadow, N., and Warner, C.M. (1998) The expression pattern of the Qa-2 antigen in murine preimplantation embryos and its correlation with the Ped gene phenotype. Mol. Human Reprod., 4, 966-971. [PDF]

Cohen, J., Scott, R., Alikani, M., Schimmel, T., Munne, S., Levron, J., Wu, L., Brenner, C., Warner, C., and Wiladsen, S. (1998) Ooplasmic transfer in mature human oocytes. Mol. Human Reprod., 4, 169-280. [PDF]

McElhinny, A.S. and Warner, C.M. (1997) Detection of cell surface protein on single preimplantation mouse embryos by Immuno-PCR. BioTechniques, 23, 660-662. [PDF]

Cai, W., Cao, W., Wu, L., Exley, G.E., Waneck, G.L., Karger, B.L., and Warner, C.M. (1996) Sequence and transcription of Qa-2 encoding genes in mouse lymphocytes and blastocysts. Immunogenetics, 45, 97-107. [PDF]

Xu, Y., Jin, P., Mellor, A.L., and Warner, C.M. (1994) Identification of the Ped gene at the molecular level: The Q9 MHC class I transgene converts the Ped slow to the Ped fast phenotype. Biol. Reprod., 51, 695-699. [PDF]

Warner, C.M., Panda, P., Almquist, C.D., and Xu, Y. (1993) Preferential survival of mice expressing the Qa-2 antigen. J. Reprod. Fertil., 99, 145-147. [PDF]

Xu, Y., Jin, P., and Warner, C.M. (1993) Modulation of preimplantation embryonic development by antisense oligonucleotides to MHC genes. Biol. Reprod., 48, 1042-1046. [PDF]

Jin, P., Meyer, T.E., and Warner, C.M. (1992) Control of embryo growth by the Ped gene: Use of reverse transcriptase-polymerase chain reaction (RT-PCR) to measure mRNA levels in preimplantation embryos. Assisted Reprod. Tech. Androl., 3, 377-383. [PDF]

Xu, Y. and Warner, C.M. (1992) Effect of antisense oligonucleotides to major histocompatibility complex (MHC) mRNA on preimplantation embryo development. Assisted Reprod. Tech. Androl., 3, 219-223. [PDF]

Tian, Z., Xu, Y., and Warner, C.M. (1992) Removal of Qa-2 antigen alters the Ped gene phenotype of preimplantation mouse embryos. Biol. Reprod., 47, 271-276. [PDF]

Warner, C.M., Brownell, M.S., and Rothschild, M.F. (1991) Analysis of litter size and weight in mice differing in Ped gene phenotype and the Q region of the H-2 complex. J. Reprod. Immunol., 19, 303-313. [PDF]

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