Basic Science Research

Enterogenesis

Short bowel syndrome, or the lack of sufficient intestine to sustain life, is a devastating clinical problem in children and adults. A number of attempts have been made to manipulate the remaining bowel to promote adaptation, and increase absorptive function. Unfortunately, none of these have been uniformly successful, and many have a high associated morbidity. The goal of this project is to use distractive forces, applied in a linear direction, to promote bowel lengthening.

Work supported by the NIH, The Hartwell Foundation and in collaboration with MC3 Corporation.


Current Results with our Pig Model:

Using a pig model, to date we have achieved a 2.7-fold increase in bowel length in 10 days. The movie shows our current hydraulic device, and experimental outcomes. Click Here For Movie

Functionally, the elongated intestine showed an increase in proliferation of epithelial cells as detected with BrdU staining.

 

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Representative histologic figures from nonoperated Jejunum, Stretch, and Control segments of pig intestine. All slides were stained for BrdU as a measure of proliferation. Note the marked increase in BrdU-positive crypt cells (brown) in the Stretch segment.

The lengthened bowel did have diminished absorptive function immediately after lengthening; however, functionality returns to normal once the bowel is re-implanted into the continuity of the pig’s intestine.

 

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Alteration of selected dissacharidase activity in Control (non-operated bowel); Lengthened bowel (immediately after lengthening); and Re-implant bowel (4 weeks after the bowel was placed into the normal intestinal continuity.

Additionally, once the lengthened bowel was re-implanted into the normal intestine, it continued to grow at a rate similar to non-operated bowel.

 

 

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Figure sustained bowel length: Note that lengthened bowel maintains its increased length, and actually continues to lengthen, after being re-implanted into the normal continuity of the pig intestine.

 

 

Collaboration with Mechanical Engineering:

Our laboratory is collaborating with Drs. Diann Brei and Jonathon Lunz in the School of Mechanical Engineering. Work in their Smart Materials Laboratory has resulted in the development of several innovative devices which drive distractive forces using shape memory alloy (SMA) activation. An example of one of these devices is shown here.

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Demonstration of the SMA-spring actuator device. A step occurs in five stages: 1) The SMA (red line) is heated resulting in contraction, pulling the collar (blue) towards the shell (Black). The spring (green) is under a bias pre-compression. 2) During contraction, the collar flap engages to the rod’s thread, while the front flap is disengaged by the thread. 3) As SMA actuation completes, both flaps engage, and the spring is fully compressed. 4) As the SMA cools, the spring force is now sufficient to push the collar back, re-extending the SMA wire. The collar flap disengages as the flap is wedged up by the thread. 5) Equilibrium between the spring and wire is reached.

 

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Picture of the SMA device as currently built. This shows the stainless steel collar pulled back along the outer shell for better visibility.


More recent designs of an implantable device for intestinal lengthening have been developed and are shown below:

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The above schematic has been manufactured, and successfully tested in a pig model.  The final device (shown below) was built by Brent Utter, and won the National Hardware Competition, in the Smart Materials Adaptive Structures and Intelligent Systems, American Society of Mechanical Engineers, 2010.

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The Teitelbaum and Brei/Luntz Laboratories were the 2011 recipients of the Ted Kennedy Family Team Excellence Award for engineering at the University of Michigan!


Recent publication by our group on enterogenesis:

pdf.jpg  Enterogenesis in a clinically feasible model of mechanical small-bowel lengthening.  Spencer AU, Sun X, El-Sawaf M, Haxhija EQ, Brei D, Luntz J, Yang H, Teitelbaum DH.  Surgery. 2006 Aug;140(2):212-20.

pdf.jpg  Luntz, J, Brei, D, Teitelbaum, DH, Spencer, AU: Mechanical extension implants for short-bowel syndrome.  Proc Soc Photo Opt Instrum Eng. Vol. 6173, p. 69-79, Smart Structures and Materials 2006: Smart Structures and Integrated Systems; Yuji Matsuzaki; Ed.

In Press: 

  1. Utter, B, Barnes, BM, Luntz, JE, Brei, D, Miysasaka, E, Okawada, M, Teitelbaum, DH: Design of an SMA Actuated Mechanotransductive Implant for Correcting Short Bowel Syndrome.  SMASIS Conference Paper, 2010.
  2. Okawada, M, Maria, HM, Teitelbaum, DH: Distraction induced enterogenesis: A unique mouse model using polyethylene glycol. J Surg Research, 2011. E-pub, PMID 21605872
  3. Miyasaka, EA, Okawada, M, Utter, B, Luntz, J, Brei, D, Teitelbaum, DH: Flow through a mechanical distraction enterogenesis device: A pilot test. J Surg Research, 2011 [Epub ahead of print].  PMID:  21571307
  4. Koga, H, Sun, X, Yang, H, Nose, K, Somara, S, Bitar, KN, Owyang, C, Okawada, M, Teitelbaum, DH: Distraction-induced intestinal enterogenesis: Preservation of intestinal function and lengthening after re-implantation into normal jejunum. Ann Surg (Accepted).


Final U.S. Patent is approved for initial enterogenesis device, May25, 2010:

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Total Parenteral Nutrition and the Mucosal Immune System

Total parenteral nutrition (TPN), or the absence of enteral nutrition, is commonly used on a clinical basis. A major consequence of patients receiving TPN is a loss of systemic and particularly mucosal immune function. This can lead to an increased incidence of infectious complications. A long-standing project in our laboratory is to understand what mechanisms contribute to these complications. One potential source of organisms entering the host is via a loss of epithelial barrier function.

We have shown that in a mouse model of TPN administration, the mucosal lymphocytes, or intraepithelial lymphocytes, increase the expression of interferon gamma (IFN-γ) and decrease the expression of IL-10. Both of these changes in cytokine expression have been shown to contribute to loss of intestinal barrier function. This loss is associated with a disruption, and internalization of several tight junction and adherens junction proteins.

 

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Immunofluorescent staining of E-cadherin (Green) and beta-catenin (red). Note in control (enterally fed mice) that these molecules are located densely along the epithelial border, and co-localize (yellow color). In the TPN group, E-cadherin moves away from the cell surface, and becomes internalized.

Our work has further shown that TPN results in a loss of epithelial cell-derived IL-7 expression, which may have a major role in the alteration of IEL phenotype and function with TPN administration.

 

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Potential model of how TPN alters epithelial cell physiology. Note that administration of TPN results in a decreased expression of IL-7, as well as an increase in IEL expression of IFN-γ and decrease in IL-10. These changes in cytokines can result in a loss of epithelial barrier function, and potentially, a loss of epithelial proliferation and apoptosis. Reversing the decline in IL-7 can reverse many of these actions.

Together, a model which may explain the mechanism of TPN associated intestinal barrier loss can be seen in the above figure.

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Our lab made the front cover of Journal of Physiology: February, 2009.

Inside the issue we demonstrate how administration of TPN results in a dissociation of cell membrane associated E-cadherin and beta catenin, with a subsequent ubiquination of beta catnenin and loss of epithelial cell proliferation.

Click here to see the abstract of this work by Yongjia Feng and Xiaoyi Sun

Front cover of Journal of Interferon and Cytokine Research: February, 2010: Glutamine prevents total parenteral nutrition-associated changes to intraepithelial lymphocyte phenotyep and function: a potential mechanism for the preservation of epithelial barrier function, Nose, et al

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In this article by first author Keisuke Nose, we demostrate potential mechanisms by which glutamine prevents the loss of epithelial barrier function with TPN administration.

Click here for the abstract.


Our lab was highlighted in the American Journal of Physiology: July, 2010, as example of translational research:
Decreased phospho-Akt signaling in a mouse model of total parenteral nutrition. A potential mechanism for the development of intestinal mucosal atrophy, Feng, et al

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The image shows changes in the abundance of mTOR with the administration of TPN (loss of functional mTOR in control TAT+TCL1G), and the ability to reverse these adverse changes in mTOR by driving pAkt using a novel TAT promoter (TPN+TCL1).

Click here for link to abstract


Collaboration with
the Department of Cellular Biology and Development and Organogenesis Center

Based on these findings, we have collaborated with Deborah Gumucio, Ph.D. in the Department of Cellular Biology and Development and the Center for Organogenesis to produce an epithelial cell IL-7 over-expressing mouse.  We hope to use this mouse to gain insight into the role of IL-7 in the pathogenesis and potential treatment of TPN-induced changes to the intestinal immune system.


Intestinal Microbiome and TPN:
Our laboratory has recently collaborated with the Huffnagle and Schloss laboratories to identify key changes in the microbiome of the small and large bowel during the administration of TPN.  454 Pyrosequencing demonstrates marked changes in the bacteria in TPN mice.

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Using MOTHUR analysis in the Schloss laboratory, principal component analysis demonstrates unique differences in the bacteria between chow fed and TPN mice, and a loss of diversity between the colon and small bowel of TPN mice.



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 Publications on IEL Epithelial Interactions with TPN:

pdf.jpg  Intestinal epithelial cell-derived interleukin-7: A mechanism for the alteration of intraepithelial lymphocytes in a mouse model of total parenteral nutrition.  Yang H, Sun X, Haxhija EQ, Teitelbaum DH.  Am J Physiol Gastrointest Liver Physiol. 2007 Jan;292(1):G84-91.

 

pdf.jpg  Interleukin-7 administration alters intestinal intraepithelial lymphocyte phenotype and function in vivo.  Yang H, Spencer AU, Teitelbaum DH.  Cytokine. 2005 Sep 21;31(6):419-28

 

pdf.jpg  Intestinal intraepithelial lymphocyte gamma delta-T cell-derived keratinocyte growth factor modulates epithelial growth in the mouse.  Yang H, Antony PA, Wildhaber BE, Teitelbaum DH.  J Immunol. 2004 Apr 1;172(7):4151-8.

 

pdf.jpg  Alteration in epithelial permeability and ion transport in a mouse model of total parenteral nutrition.  Yang H, Finaly R, Teitelbaum DH.  Crit Care Med. 2003 Apr;31(4):1118-25.

pdf.jpg  Decline in Intestinal Mucosal IL-10 Expression and Decreased Intestinal Barrier Function in a Mouse Model of Total Parenteral Nutrition.  Sun X, Yang H, Nose K, Nose S, Haxhija EQ, Koga H, Feng Y, Teitelbaum DH.  Am J Physiol Gastrointest Liver Physiol. 2008 Jan;294(1):G139-47

pdf.jpg Intestinal specific overexpression of interleukin-7 attenuates the alternation of intestinal intraepithelial lymphocytes after total parenteral nutrition administration.  Yang H, Gumucio DL, Teitelbaum DH.  Ann Surg. 2008 Nov;248(5):849-56.

pdf.jpg Specific overexpression of IL-7 in the intestinal mucosa: the role in intestinal intraepithelial lymphocyte development.  Yang H, Madison B, Gumucio DL, Teitelbaum DH.  Am J Physiol Gastrointest Liver Physiol. 2008 Jun;294(6):G1421-30

pdf.jpg Dissociation of E-Cadherin and {beta}-catenin in a mouse model of total parenteral nutrition: A mechanism for the loss of epithelial cell proliferation and villus atrophy.  Feng Y, Sun X, Yang H, Teitelbaum DH.  J Physiol. 2009, 587(Pt 3):641-54.



Blockade of the Renin-Angiotensin System in the Intestinal Mucosa and Inflammatory Bowel Disease

Our laboratory has found that the expression of angiotensin converting enzyme in the intestinal mucosa is highly correlated to epithelial cell apoptosis. Use of enalaprilat, an angiotensin converting enzyme inhibitor (ACE-I), can markedly down-regulate epithelial cell apoptosis, and decrease the local expression of tumor necrosis factor alpha (TNF-α).

Based on these findings, we hypothesized that the use of enalaprilat could prevent or decrease many of the changes observed in models of inflammatory bowel disease. Our laboratory has recently developed a compound which allows us to deliver an ACE-I to the colonic mucosa, while preventing systemic absorption of the inhibitor, and resultant hypotension. Use of this agent resulted in a significant reduction in the histologic findings of a mouse dextran sodium sulfate model

 

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ACE-I histologic changes in DSS model Micrographic examples of colon in dextran sodium sulfate model treated with enalaprilat or placebo. Note the markedly improved architecture of the mucosa. As well, the brown stained cells are TUNEL positive cells, indicating apoptosis. Note the decreased number of apoptotic cells in the enalaprilat treated mice.

 

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TNF expression TNF-a expression (via real time PCR) was markedly decreased in DSS-treated mice given enalaprilat in a dose-dependent fashion. *P<0.05.

 


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Enalaprilat PEG movie.  Development of a new compound for inflammatory bowel disease

Click Here for Movie Demonstration

Collaboration with the Department of Medicinal Chemistry, School of Pharmacology

Based on these findings, we have collaborated with Hollis Showalter, Ph.D. and Doug Larsen, Ph.D. in the Department of Medicinal Chemistry and their Medicinal Chemistry Core Synthesis Laboratory, and Peter Lucas in Pathology to work on the production of a whole new class of agents which can block the Renin-Angiotensin System at the level of the Angiotensin II type 1a (AT1a) receptor. 


A demonstration of the efficacy of blocking AT1a receptors is shown below for three such antagonists, losartan, deschlorolosartan (DCL) which lacks systemic exposure, and candesartan.  Note the dramatic prevention of colitis in the treated groups, and reduction in the development of apoptosis (TUNEL staining, right panels).

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 The Teitelbaum Laboratory is currently collaborating with TSRL Inc, Ann Arbor, MI, via two SBIR small business grants.

 Recent publications by our group:


pdf.jpg  Intestinal intraepithelial lymphocyte derived angiotensin converting enzyme modulates epithelial cell apoptosis.   Wildhaber BE, Yang H, Haxhija EQ, Spencer AU, Teitelbaum DH.  Apoptosis. 2005 Dec;10(6):1305-15.

pdf.jpg  Reduced severity of a mouse colitis model with angiotensin converting enzyme inhibition.   Spencer AU, Yang H, Haxhija EQ, Wildhaber BE, Greenson JK, Teitelbaum DH.  Dig Dis Sci. 2007 Apr;52(4):1060-70.

The role of angiotensin II type 1a receptor on intestinal epithelial cells following small bowel resection in a mouse model.  Koga H, Yang H, Haxhija EQ, Teitelbaum DH.  Pediatr Surg Int. 2008 Dec;24(12):1279-86

Transanal delivery of angiotensin converting enzyme inhibitor prevents colonic fibrosis in a mouse colitis model: development of a unique mode of treatment.   Koga H, Yang H, Adler J, Zimmermann EM, Teitelbaum DH.      Surgery. 2008 Aug;144(2):259-68.

Modulation of mouse intestinal epithelial cell turnover in the absence of angiotensin converting enzyme.  Haxhija EQ, Yang H, Spencer AU, Koga H, Sun X, Teitelbaum DH. Am J Physiol Gastrointest Liver Physiol. 2008 Jul;295(1):G88-G98.

Use of Enterally Delivered Angiotensin II Type Ia Receptor Antagonists to Reduce the Severity of Colitis.  Okawada, Koga H, Larsen SD, Showalter HD, Turbiak AJ, Jin X, Lucas PC, Lipka E, Hillfinger J, Kim JS, Teitelbaum DH.: Dig Dis Sci. 2011 Mar 12. [Epub ahead of print].  PMID:  21399927

Creation of a Human Bioartificial Internal Anal Sphincter

    The GI Molecular Motors lab run by Dr. Khalil Bitar and our laboratory were just awarded a NIH Research Challenge Grant to further our work in the development of a bioartificial internal anal sphincter for the treatment of fecal incontinence. 
    Recent publications by our two laboratories:

Hashish M, Raghavan S, Somara S, Gilmont RR, Miyasaka E, Bitar KN, Teitelbaum DH.: Surgical implantation of a bioengineered interanal anal sphincter. J Pediatr Surgery, 2010, 45(1):52-8.

Raghavan S, Miyasaka EA, Hashish M, Somara S, Gilmont RR, Teitelbaum DH, Bitar KN.: Successful implantation of physiologically functional bioengineered mouse internal anal sphincter.  Am J Physiol Gastrointest Liver Physiol. 2010 Aug;299(2):G430-9

Miyasaka EA, Raghavan S, Gilmont RR, Mittal K, Somara S, Bitar KN, Teitelbaum DH.:In vivo growth of a bioengineered internal anal sphincter: comparison of growth factors for optimization of growth and survival.  Pediatr Surg Int. 2010 Nov 3. [Epub ahead of print]


The images below shows the formation of an internal anal sphincter (IAS) grown from harvested human internal anal sphincter, and its successful reimplantation into a nude mouse (3 weeks post-implantation and treatment with FGF-2).  Also shown are the method for creating this IAS.

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