Watercress-based electrospun nanofibrous scaffolds enhance proliferation and stemness preservation of human adipose-derived stem cells

Figures & data

Table 1. Description of primer sets used in qPCR for gene expression analyses.

Gene	Accession no.	Primers 5′ → 3′	PCR product (bp)
Sox-2	NM_003106	F: TTACCTCTTCCT CCC ACT CCA R: GGTAGTGCTGGGACATGTGAA	132
Nanog	NM_024865	F: CTGTGATTTGTGGGCCTGAA R: TGTTTGCCTTTGGGACTGGT	153
Rex-1	NM_174900	F: AAAGGTTTTCGAAGCAAGCTC R: CTGCGAGCTGTT TAG GAT CTG	185
Oct-4	NM_002701	F: AAGGATGTGGTCCGAGTGTG R: GAAGTGAGGGCTCCCATAGC	180
Cyclin D1	NM_053056	F: AGACCTTCGTTGCCCTCTGT R:CAGTCCGGGTCACACTTGAT	181
pRb	NM_000321	F: CAGACCCAGAAGCCATTGAA R:CTGGGTGCTCAGACAGAAGG	115
p53	NM_001126112.2	F: CCCAGCCAAAGAAGAAACCA R:GTTCCAAGGCCTCATTCAGCT	101
GAPDH	NM_002046	F: TCCCTGAGCTGAACGGGAAG R: GGAGGAGTGGGTGTCGCTGT	217

Table 2. Antioxidant activity of WE-loaded PCL/PEG nanofibers (n = 3).

Optical density value (OD515nm ± SD)
Time /min	WE-loaded PCL/PEG	Control sample	AA%
0	1.875 ± 0.08	1.976 ± 0.07	2
30	1.654 ± 0.12	1.962 ± 0.05	16.34
60	1.512 ± 0.08	1.978 ± 0.07	23.80
90	1.222 ± 0.11	1.984 ± 0.11	37.62

Figure 1. SEM images of fabricated nanofibers, PCL-PEG (A), WE-PCL-PEG (B and C).

Figure 2. FTIR spectra of WE, PCL-PEG and WE-PCL-PEG nanofibers.

Figure 3. Cell viability of ADSCs grown PCL-PEG nanofibers, WE-PCL-PEG nanofibers and tissue culture plates (TCPs) after 1, 7 and 14 days of cell culture. *p < .05. Values are expressed as mean ± SD of three parallel measurements.

Figure 4. FE-SEM images of ADSCs morphology on the PCL-PEG nanofibers (A, B, C) and WE-loaded PCL-PEG nanofibers (D, E, F) after 7 and 14 days of seeding.

Figure 5. Viability of ADSCs cultured in the presence and absence of 1% H₂O₂ on PCL-PEG nanofibers and WE-loaded PCL-PEG nanofibers. Viability was significantly reduced on PCL-PEG nanofibers lacking WE due to oxidative stress (* p ≤ 0.05). Results are mean ± SD (n = 3).

Figure 6. DNA content of of ADSCs cultured on control PCL-PEG nanofibers (A) and WE-PCL-PEG nanofibers (B). Results are mean ± SD (n = 3).

Table 3. Percentage of cells at various cell phases (values are expressed in mean ± SD).

	Cell cycle phase
Group	G0/G1%	S %	G2/M%	PI %
PCL/PEG	84/31 ± 0.006	9/57 ± 0.004	6/12 ± 0.003	15.67 ± 0.228
WE-PCL/PEG	82/35 ± 0.009	13/57 ± 0.007	3/95 ± 0.006	17.52±.561

Figure 7. The expression levels of cell cycle–regulated genes (Cyclin D1, pRb and p53) were investigated by qRT-PCR in ADSCs cultured on PCL-PEG nanofibers, WE-PCL-PEG nanofibers and control (without scaffold) for 7 and 14 days. The cell cycle–regulated genes were significantly up-regulated in WE-loaded PCL-PEG nanofibers as compared to PCL-PEG (*p < .05). Results are mean ± SD (n = 3).

Figure 8. The expression of Oct-4, Rex-1, Sox-2 and Nanog3 genes were investigated by qRT-PCR in cultured on PCL-PEG scaffold, WE-PCL-PEG and control (without scaffold). GAPDH gene was used as internal control. All stemness marker genes, were significantly up-regulated in WE-loaded PCL-PEG nanofibers as compared to PCL-PEG (* p < .05). Results are mean ± SD (n = 3).