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Beneficial reciprocal effects of bone marrow stromal cells and Schwann cells from adult rats in a dynamic co‑culture system in vitro without intercellular contact.

Zhou LN, Cui XJ, Su KX, Wang XH, Guo JH - Mol Med Rep (2015)

Bottom Line: In order to examine how implanted bone marrow stromal cells (BMSCs) encourage peripheral nerve regeneration, the present study investigated the interaction of BMSCs and Schwann cells (SCs) using an indirect in vitro co‑culture model.On the 3rd day after co‑culture, only a few co‑cultured BMSCs showed the typical SC‑like morphology, while most BMSCs still kept their native appearance.These results indicated that BMSCs may interact synergistically with SCs with regard to promoting peripheral nerve regeneration.

View Article: PubMed Central - PubMed

Affiliation: Department of Anatomy, Guangdong Medical University, Zhanjiang, Guangdong 524023, P.R. China.

ABSTRACT
In order to examine how implanted bone marrow stromal cells (BMSCs) encourage peripheral nerve regeneration, the present study investigated the interaction of BMSCs and Schwann cells (SCs) using an indirect in vitro co‑culture model. SCs and BMSCs were obtained from adult Sprague‑Dawley rats. The passaged BMSCs were CD29‑ and CD44‑positive but CD45‑negative and were co‑cultured with the primary SCs using a Millicell system, which allows BMSCs and SCs to grow in the same culture medium but without direct contact. Expression of the typical SC markers S‑100 and glial fibrillary acidic protein (GFAP) of the treated BMSCs as well as the proliferation capacity of the co‑cultured SCs was evaluated by immunocytochemical staining on the 3rd and 5th day of co‑culture. Immunocytochemical staining showed that >75% of the BMSCs in the indirect co‑culture model were GFAP‑ and S‑100‑positive on the 3rd and 5th day after co‑culture, as opposed to <5% of the BMSCs in the control group. On the 3rd day after co‑culture, only a few co‑cultured BMSCs showed the typical SC‑like morphology, while most BMSCs still kept their native appearance. By contrast, on the 5th day after co‑culture, almost all of the co‑cultured BMSCs appeared with the typical SC‑like morphology. Furthermore, 70.71% of the SCs in the indirect co‑culture model were S‑100‑positive on the 5th day of co‑culture, as opposed to >30.43% of the SCs in the control group. These results indicated that BMSCs may interact synergistically with SCs with regard to promoting peripheral nerve regeneration.

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Induction of SC proliferation by diffusible factors of BMSCs. The proliferation of SCs co-cultured with (A–C) BMSCs and (D–F) control was assessed by (A and D) S-100 (red) immunofluorescence and (B and E) Hoechst 33258 nuclear staining (blue) (scale bar, 100 µm). (G) The number of S-100-positive cells as a percentage of the number of Hoechst-positive nuclei was averaged from three wells and repeated four times. Each of the four repeats was from a different culture, with each culture in turn being derived from a different animal. Quantitative analysis showed that the number of SCs co-cultured with BMSCs (70.71±1.24%, n=6) was significantly higher than that of the control SCs (30.43±2.23%, n=6) (*P<0.05). BMSC, bone marrow stromal cell; SC, Schwann cell.
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f2-mmr-12-04-4931: Induction of SC proliferation by diffusible factors of BMSCs. The proliferation of SCs co-cultured with (A–C) BMSCs and (D–F) control was assessed by (A and D) S-100 (red) immunofluorescence and (B and E) Hoechst 33258 nuclear staining (blue) (scale bar, 100 µm). (G) The number of S-100-positive cells as a percentage of the number of Hoechst-positive nuclei was averaged from three wells and repeated four times. Each of the four repeats was from a different culture, with each culture in turn being derived from a different animal. Quantitative analysis showed that the number of SCs co-cultured with BMSCs (70.71±1.24%, n=6) was significantly higher than that of the control SCs (30.43±2.23%, n=6) (*P<0.05). BMSC, bone marrow stromal cell; SC, Schwann cell.

Mentions: As shown by double labelling with S-100 (Fig. 2A) and Hoechst 33258 (Fig. 2B) on the fifth day of co-culture, SCs co-cultured with BMSCs exhibited a clear spindle-shaped or triangular soma with two or three slender processes (Fig. 2A and C). Either S-100-labelled cells (Fig. 2A and D) or Hoechst 33258-labelled nuclei (Fig. 2B and E) were obviously more numerous in the BMSCs co-culture system than in the acellular control cultures (Fig. 2C and F). The quantitative study showed that the number of S-100-positive SCs as a percentage of the number of Hoechst-labelled nuclei in the BMSC co-culture was significantly higher than that in the acellular control culture, after 5 days (70.71±1.24 vs. 30.43±2.23%; P<0.05) (Fig. 2G).


Beneficial reciprocal effects of bone marrow stromal cells and Schwann cells from adult rats in a dynamic co‑culture system in vitro without intercellular contact.

Zhou LN, Cui XJ, Su KX, Wang XH, Guo JH - Mol Med Rep (2015)

Induction of SC proliferation by diffusible factors of BMSCs. The proliferation of SCs co-cultured with (A–C) BMSCs and (D–F) control was assessed by (A and D) S-100 (red) immunofluorescence and (B and E) Hoechst 33258 nuclear staining (blue) (scale bar, 100 µm). (G) The number of S-100-positive cells as a percentage of the number of Hoechst-positive nuclei was averaged from three wells and repeated four times. Each of the four repeats was from a different culture, with each culture in turn being derived from a different animal. Quantitative analysis showed that the number of SCs co-cultured with BMSCs (70.71±1.24%, n=6) was significantly higher than that of the control SCs (30.43±2.23%, n=6) (*P<0.05). BMSC, bone marrow stromal cell; SC, Schwann cell.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4581791&req=5

f2-mmr-12-04-4931: Induction of SC proliferation by diffusible factors of BMSCs. The proliferation of SCs co-cultured with (A–C) BMSCs and (D–F) control was assessed by (A and D) S-100 (red) immunofluorescence and (B and E) Hoechst 33258 nuclear staining (blue) (scale bar, 100 µm). (G) The number of S-100-positive cells as a percentage of the number of Hoechst-positive nuclei was averaged from three wells and repeated four times. Each of the four repeats was from a different culture, with each culture in turn being derived from a different animal. Quantitative analysis showed that the number of SCs co-cultured with BMSCs (70.71±1.24%, n=6) was significantly higher than that of the control SCs (30.43±2.23%, n=6) (*P<0.05). BMSC, bone marrow stromal cell; SC, Schwann cell.
Mentions: As shown by double labelling with S-100 (Fig. 2A) and Hoechst 33258 (Fig. 2B) on the fifth day of co-culture, SCs co-cultured with BMSCs exhibited a clear spindle-shaped or triangular soma with two or three slender processes (Fig. 2A and C). Either S-100-labelled cells (Fig. 2A and D) or Hoechst 33258-labelled nuclei (Fig. 2B and E) were obviously more numerous in the BMSCs co-culture system than in the acellular control cultures (Fig. 2C and F). The quantitative study showed that the number of S-100-positive SCs as a percentage of the number of Hoechst-labelled nuclei in the BMSC co-culture was significantly higher than that in the acellular control culture, after 5 days (70.71±1.24 vs. 30.43±2.23%; P<0.05) (Fig. 2G).

Bottom Line: In order to examine how implanted bone marrow stromal cells (BMSCs) encourage peripheral nerve regeneration, the present study investigated the interaction of BMSCs and Schwann cells (SCs) using an indirect in vitro co‑culture model.On the 3rd day after co‑culture, only a few co‑cultured BMSCs showed the typical SC‑like morphology, while most BMSCs still kept their native appearance.These results indicated that BMSCs may interact synergistically with SCs with regard to promoting peripheral nerve regeneration.

View Article: PubMed Central - PubMed

Affiliation: Department of Anatomy, Guangdong Medical University, Zhanjiang, Guangdong 524023, P.R. China.

ABSTRACT
In order to examine how implanted bone marrow stromal cells (BMSCs) encourage peripheral nerve regeneration, the present study investigated the interaction of BMSCs and Schwann cells (SCs) using an indirect in vitro co‑culture model. SCs and BMSCs were obtained from adult Sprague‑Dawley rats. The passaged BMSCs were CD29‑ and CD44‑positive but CD45‑negative and were co‑cultured with the primary SCs using a Millicell system, which allows BMSCs and SCs to grow in the same culture medium but without direct contact. Expression of the typical SC markers S‑100 and glial fibrillary acidic protein (GFAP) of the treated BMSCs as well as the proliferation capacity of the co‑cultured SCs was evaluated by immunocytochemical staining on the 3rd and 5th day of co‑culture. Immunocytochemical staining showed that >75% of the BMSCs in the indirect co‑culture model were GFAP‑ and S‑100‑positive on the 3rd and 5th day after co‑culture, as opposed to <5% of the BMSCs in the control group. On the 3rd day after co‑culture, only a few co‑cultured BMSCs showed the typical SC‑like morphology, while most BMSCs still kept their native appearance. By contrast, on the 5th day after co‑culture, almost all of the co‑cultured BMSCs appeared with the typical SC‑like morphology. Furthermore, 70.71% of the SCs in the indirect co‑culture model were S‑100‑positive on the 5th day of co‑culture, as opposed to >30.43% of the SCs in the control group. These results indicated that BMSCs may interact synergistically with SCs with regard to promoting peripheral nerve regeneration.

Show MeSH
Related in: MedlinePlus