Obstet Gynecol Sci.  2020 Sep;63(5):594-604. 10.5468/ogs.20073.

Isolation of mesenchymal stem cells from Pap smear samples

• Park J
• Son D
• Hong W
• Jang J
• Cho GJ
• Song G
• Kim IY
• You S

• KMID: 2506502
• DOI: http://doi.org/10.5468/ogs.20073

Abstract

Objective
Exploiting their ability to differentiate into mesenchymal lineages like cartilage, bone, fat, and muscle, and to elicit paracrine effects, mesenchymal stem cells (MSCs) are widely used in clinical settings to treat tissue injuries and autoimmune disorders. One of accessible sources of MSC is the samples used for Papanicolaou (Pap) test, which is a cervical screening method for detecting potentially pre-cancerous and cancerous alterations in the cervical cells and to diagnose genetic abnormalities in fetuses. This study aimed to identify and isolate the stem cells from Pap smear samples collected from pregnant women, and to trace the origin of these cells to maternal or fetal tissue, and characterize their stem cell properties.
Methods
To investigate the possibility and efficiency of establishing MSC lines from the Pap smear samples, we were able to establish 6 cell lines from Pap smear samples from 60 pregnant women at different stages of gestation.
Results
The 3 cell lines randomly selected among the 6 established in this study, displayed high proliferation rates, several characteristics of MSCs, and the capacity to differentiate into adipocytes, osteocytes, and chondrocytes. Our study identified that the stem cell lines obtainable from Pap smear sampling were uterine cervical stromal cells (UCSCs) and had 10% efficiency of establishment.
Conclusion
Despite their low efficiency of establishment, human UCSCs from Pap smear samples can become a simple, safe, low-cost, and donor-specific source of MSCs for stem cell therapy and regenerative medicine.

Keyword

Papanicolaou test; Mesenchymal stem cells; Regenerative medicine

Figure

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  Fig. 1. Isolation of fibroblast-like cells from Papanicolaou (Pap) smear samples. (A) Method used to isolate cells from Pap smear samples. (B) Morphology of isolated cell line at passage 0 and 5 (scale bar=500 µm). (C) Accumulated population comparison with isolated cell line and amniotic fluid-derived mesenchymal stem cell (AF-MSC) upon passaging. (D) Immunofluorescence staining and fluorescence-activated cell sorting (FACS) analysis of human leukocyte antigen (HLA)-G in AF-MSCs and isolated cells (scale bar=200 μm). (E) Karyotype of isolated cell line determined by G-banding.

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  Fig. 2. Characterization of human uterine cervical stromal cell (hUCSC) lines. (A) Accumulated population of hUCSCs and amniotic fluidderived mesenchymal stem cell (AF-MSC) upon passaging. (B) Immunofluorescence staining of human leukocyte antigen (HLA)-G in hUCSC line #2 and #3 (scale bar=200 μm). Fluorescence-activated cell sorting (FACS) analysis of HLA-G expression in hUCSC line #2 and #3. The percentages of HLA-G-positive cells are shown. (C) Karyotype of hUCSC line #2 and #3 determined by G-banding. (D) FACS analysis of the immunophenotypes of hUCSC lines (green). The isotype control is shown in purple. (E) mRNA expression of human MSC markers in hUCSC lines. (F) Colony-forming unit (CFU) assay investigating the self-renewal capacity of hUCSC lines.

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  Fig. 3. Differentiation of human uterine cervical stromal cells (hUCSCs). (A) Adipogenic differentiation of hUCSCs at passage 8. Differentiation was evaluated by Oil Red O staining and reverse transcription polymerase chain reaction (RT-PCR) analysis of adipocyte markers (LPL and aP2). (B) Osteogenic differentiation of hUCSCs at passage 8. Differentiation was evaluated by von Kossa staining and RT-PCR analysis of osteocyte markers (osteopontin and osteocalcin). (C) Chondrogenic differentiation of hUCSCs at passage 7. Differentiation was evaluated by Alcian Blue staining and RT-PCR analysis of chondrocyte markers (collagen I and aggrecan). Scale bar=200 µm. AF-MSC, amniotic fluid-derived mesenchymal stem cell; GAPDH, glyceraldehyde 3-phosphate dehydrogenase.

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  Fig. 4. Establishment of human uterine cervical stromal cells (hUCSCs) from Papanicolaou (Pap) smear samples. MSC, mesenchymal stem cell.

Reference

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