Korean J Urol.  2009 May;50(5):480-485.

The Effect of Human Muscle-Derived Stem Cells (MDSC) and Glycine-Isoleucine-Lysine-Valine-Alanine-Valine (GIKVAV) on the Cryo-Injured Bladder of Nude Mouse

Affiliations
  • 1Department of Urology, The Catholic University of Korea College of Medicine, Seoul, Korea. uroljy@catholic.ac.kr
  • 2Department of Urology, Seoul National University College of Medicine, Seoul, Korea.
  • 3Department of Biomaterials Research Center, Korea Institute of Science and Technology, Seoul, Korea.

Abstract

PURPOSE: In neurogenic bladder, both smooth muscle contraction and nerve regeneration are very important for functional improvement. Glycine-isoleucine-lysine-valine-alanine-valine (GIKVAV) is a peptide that can induce nerve regeneration in vivo. In this study, we evaluated bladder function after injection of muscle-derived stem cells (MDSCs) and GIKVAV into the cryo-injured bladder of nude mice.
MATERIALS AND METHODS
Human muscle samples were obtained from the rectus abdominis muscle of 12 patients who underwent laparotomy. The purpose and entire method of the study were explained to the patients, and all subjects who participated in this study provided written informed consent. The MDSCs were isolated by a modified preplate technique, and only CD34+ human MDSC were extracted by use of Mini-MACS kits. The nude mice were subdivided into 5 groups (n=40): normal group (N, n=8), saline injection group after cryo-injury (S, n=8), GIKVAV injection group after cryo-injury (G, n=8), human MDSC injection group after cryo-injury (M, n=8), and GIKVAV and human MDSC injection group after cryo-injury (GM, n=8). At 2 weeks after injection, we compared the contractility of a bladder muscle strip of each group by organ bath and polygraph by using electronic field stimulation (EFS). Nerve regeneration was evaluated by choline acetyl transferase (ChAT) immunostaining.
RESULTS
The contractile powers of the N, S, G, M, and GM groups were 3.58+/-0.27, 1.54+/-0.25, 1.54+/-0.31, 2.49+/-0.36, and 2.44+/-0.34 mN/mg, respectively, by EFS. The contractility of the bladder muscle strip in the S and G groups was lower than that in the N group. The contractile powers of the M and GM groups were lower than those of the N group but greater than those of the S and G groups. In ChAT immunohistochemical staining, nerve regeneration was increased in the G and GM groups compared with the S and M groups.
CONCLUSIONS
Nerve regeneration was induced by GIKVAV injection regardless of human MDSC injection. There was no direct effect of GIKVAV on bladder muscle contractility.

Keyword

Muscles; Stem cells; Nude mice; Nerve regeneration

MeSH Terms

Animals
Baths
Choline
Contracts
Electronics
Electrons
Humans
Informed Consent
Laparotomy
Mice
Mice, Nude
Muscle, Smooth
Muscles
Nerve Regeneration
Rectus Abdominis
Stem Cells
Transferases
Urinary Bladder
Urinary Bladder, Neurogenic
Choline
Transferases

Figure

  • Fig. 1 The organ bath study of bladder contractility by use of electrical field stimulation. The average contractile power of the bladder muscle strip in all experimental groups (S, G, M, GM) was significantly less than in the normal control group (N). The average contractile power of the M and GM groups was significantly higher than that of the S and G groups. N: normal, N group, S: saline, S group, G: glycine-isoleucine-lysine-valine-alanine-valine, G group, M: muscle-derived stem cells, M group, GM: GIKVAV+MDSC, GM group, a: p<0.01 compared with S and G groups.

  • Fig. 2 H&E and fluorescent imaging of nude mouse bladder. The black arrows show the smooth muscle thickness of each group. The bladder smooth muscle layers of the S and G groups were thinner than those of the N group (A-C, ×100). The bladder smooth muscle layers of the M and GM groups were thicker than those of the S and G groups (D, E, ×100). Newly developed muscle layers were found in the M and GM groups. In fluorescent microscopy, PKH-labeled (red colored) injected cells were observed in the newly developed smooth muscle layers in the M and GM groups (I, J, ×100) but not in the N, S, and G groups (F-H, ×100). N: normal, N group, S: saline, S group, G: glycine-isoleucine-lysine-valine-alanine-valine, G group, M: muscle-derived stem cells, M group, GM: GIKVAV+MDSC, GM group.

  • Fig. 3 Immunohistochemical staining of choline acetyl transferase in nude mouse bladder. The nuclei were colored with DAPI (blue color). Four weeks after cryo-injury, immunofluorescence analysis showed a decreased expression (green color) of choline acetyltransferase in the S and M groups compared with the N group (A, B, D, ×100). After injection of GIKAV, the expression of choline acetyl transferase (green color) was increased in the G and GM groups compared with the S and M groups (C, E, ×100). N: normal, N group, S: saline, S group, G: glycine-isoleucine-lysine-valine-alanine-valine, G group, M: muscle-derived stem cells, M group, GM; GIKVAV+MDSC, GM group.


Reference

1. Koh JS, Lee JY, Lee JY. The effects of human muscle derived stem cells on the Induction of peripheral nerve regeneration. Korean J Urol. 2008. 49:350–359.
2. Lee HN, Lee JY, Koh JS, Kim HW, Byun SS, Lee SS, et al. Muscle derived stem celll/alginate/polycaprolactone/ injection therapy in rats with denervated urethral sphincter. Korean J Urol. 2007. 48:1296–1301.
3. Choo GY, Lee JY, Park WH, Jung YS. Effects of injection therapy using muscle derived stem cell/chitosan/hydroapatite composite gel in a rat model of urinary incontinence. Korean J Urol. 2007. 48:627–632.
4. Byun SS, Chung YS, Lee SS, Lee HN, Lee JY, Lee JY. Augmentation cystoplasty using hydroxapatite/chitosan composite sheet seeded with autologous muscle-derived stem cells. Korean J Urol. 2007. 48:433–438.
5. Lin X, Takahashi K, Liu Y, Zamora PO. Enhancement of cell attachment and tissue integration by a IKVAV containing multi-domain peptide. Biochim Biophys Acta. 2006. 1760:1403–1410.
6. Qu Z, Balkir L, van Deutekom JC, Robbins PD, Pruchnic R, Huard J. Development of approaches to improve cell survival in myoblast transfer therapy. J Cell Biol. 1998. 142:1257–1267.
7. Reali C, Scintu F, Pillai R, Cabras S, Argiolu F, Ristaldi MS, et al. Differentiation of human adult CD34+ stem cells into cells with a neural phenotype: role of astrocytes. Exp Neurol. 2006. 197:399–340.
8. Somogyi GT, Yokoyama T, Szell EA, Smith CP, de Groat WC, Huard J, et al. Effect of cryoinjury on the contractile parameters of bladder strips: a model of impaired detrusor contractility. Brain Res Bull. 2002. 59:23–28.
9. Cannon TW, Lee JY, Somogyi G, Pruchnic R, Smith CP, Huard J, et al. Improved sphincter contractility after allogenic muscle-derived progenitor cell injection into the denervated rat urethra. Urology. 2003. 62:958–963.
10. Lee JY, Cannon TW, Pruchnic R, Fraser MO, Huard J, Chancellor MB. The effects of periurethral muscle-derived stem cell injection on leak point pressure in a rat model of stress urinary incontinence. Int Urogynecol J Pelvic Floor Dysfunct. 2003. 14:31–37.
11. Huard J, Yokoyama T, Pruchnic R, Qu Z, Li Y, Lee JY, et al. Muscle-derived cell-mediated ex vivo gene therapy for urological dysfunction. Gene Ther. 2002. 9:1617–1626.
12. Gunn JW, Turner SD, Mann BK. Adhesive and mechanical properties of hydrogels influence neurite extension. J Biomed Mater Res A. 2005. 72:91–97.
13. Tysseling-Mattiace MV, Sahni V, Niece KL, Birch D, Czeisler C, Fehlings MG, et al. Self-assembling nanofibers inhibit glial scar formation and promote axon elongation after spinal cord injury. J Neurosci. 2008. 28:3814–3823.
Full Text Links
  • KJU
Actions
Cited
CITED
export Copy
Close
Share
  • Twitter
  • Facebook
Similar articles
Copyright © 2024 by Korean Association of Medical Journal Editors. All rights reserved.     E-mail: koreamed@kamje.or.kr