Adenovirus-Mediated Antisense Vector-Induced Inhibition of Human Telomerase RNA May Induce Differentiation of CD34+ Cells

Seok Jin Kim; Joon-Seok Song; Chang Hee Song; Ji Hyun Yoo; Byung Soo Kim

doi:10.5045/kjh.2006.41.3.172

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Kim, Song, Song, Yoo, and Kim: Adenovirus-Mediated Antisense Vector-Induced Inhibition of Human Telomerase RNA May Induce Differentiation of CD34+ Cells

Original Article

Korean J Hematol 2006;41(3):172-178.

Published online: 19 September 2006

DOI: https://doi.org/10.5045/kjh.2006.41.3.172

Adenovirus-Mediated Antisense Vector-Induced Inhibition of Human Telomerase RNA May Induce Differentiation of CD34+ Cells

Seok Jin Kim, M.D., Ph.D., Joon-Seok Song, Ph.D., Chang Hee Song, Ji Hyun Yoo, Byung Soo Kim, M.D., Ph.D.

Department of Internal Medicine, Korea University Medical Center, Instititue of Stem Cell Research, Korea University, Seoul, Korea

Correspondence to：Byung Soo Kim, M.D., Ph.D. Division of Oncology and Hematology, Department of Internal Medicine, Korea University Medical Center 126-1, Anam-dong 5-ga, Seongbuk-gu, Seoul 136-705, Korea Tel: +82-2-920-5980, Fax: +82-2-920-6520 E-mail: kbs0309@paran.com

This study was supported by a grant (#SC2220) from the Stem Cell Research Center of the 21^st Century Frontier Research Program funded by Ministry of Science and Technology, Republic of Korea.

Received 11 April 2006 Revised 22 August 2006 Accepted 12 September 2006

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Background:

Background Telomerase activation and human telomerase RNA (hTR) expression are known to be related to the preservation of the “stemness” of stem cells. In this study, we have inhibited the expression of hTR to find the relationship between the telomerase activity and differentiation of normal hematopoietic stem cells.

Methods:

We used cord blood collected from 10 full term pregnant women. We classified the CD34+ hematopoietic stem cells from the same donor into three groups: the Ad-OA group was treated with the recombinant adenoviral (Ad) vector Ad-OA using telomerase antisense, the Ad-M6 group was treated with a mutant version of the Ad-OA without telomerase antisense, and a control group without any treatment.

Results:

The mean number of colony-forming cells (CFCs) were 110±38 for the Ad-OA groups, 540±56 for the Ad-M6 groups, and 650±72 for the control groups. Thus, CFCs in the Ad-OA group were lower than in the Ad-M6 group (P＜0.01). The myeloid portion of the CFCs in the Ad-OA group was higher than the Ad-M6 and control groups (P＜0.01). The Ad-OA group showed a higher percentage of granulocytes suggesting more of a tendency for myeloid differentiation than the Ad-M6 and control groups (P＜0.01). We found that the suppression of telomerase activity by the antisense telomerase adenovirus induced the differentiation of hematopoietic stem cells confirmed by differential cell count and cytochemical staining.

Conclusion:

These findings suggest that the activity of the telomerase may play a role in the differentiation of normal CD34+ hematopoietic stem cells into mature cells.

Keywords: Telomerase, Antisense adenovirus, Hematopoietic stem cell, Differentiation

REFERENCES

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Fig. 1

Structure of recombinant adenoviral vector with deficient E1a/E1b. It contains residues 94 and 76 of the telomerase template sequence driven by the cytomegalovirus immediate promoter, inserted into the E1 region.

Fig. 2

Cell morphology in Ad-OA and Ad-M6 groups at day 7 of long-term culture with CD34+ cells. Top. Blood cell morphology with Wright Stain (×400): significantly higher percentage of granulocytes in the Ad-OA group compared to (B) those in the Ad-M6 group (A) this suggests a differentiation induction effect of telomerase antisense. Bottom. Granulocyte morphology of the myeloperoxidase stain (×1,000): There is a more dense myelopeoxidase stain pattern in the Ad-OA group when compared to (D) that in the Ad-M6 group (C) this suggests a maturation induction effect of telomerase antisense.

Fig. 3

The degree of apoptosis in CD34+ cells of Ad-OA, Ad-M6, and control Mock groups. The data represent mean values and standard deviation (S.D.) of three independent experiments.

Table 1.

Differential cell counts of Ad-OA, Ad-M6, and control group a3 and 7 day after long-term culture with CD34+ cells

	Day 3 (%, mean±SEM)			Day 7 (%, mean±SEM)
	Ad-OA	Ad-M6	Control	Ad-OA	Ad-M6	Control
Blast	21±6	47±12	68±8	2±1	18±9	20±6
Lymphocyte	17±7	16±7	18±7	8±3	5±2	16±11
Granulocyte	45±5∗	12±6	7±3	33±12∗	9±3	16±5

∗P＜0.01. Abbreviations: Ad-OA, group treated with the recombinant adenoviral (Ad) vector Ad-OA with telomerase antisense; Ad-M6, group with a mutant version of Ad-OA without telomerase antisense; control, group without any treatment.

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