Immunomodulatory effect of Berberis vulgaris extracts on murine splenocytes and enrichment of dendritic cells in vitro

Figures & data

Table 1. Primers' sequences, annealing temperatures and base pairs, corresponding to the PCR product.

Primers' sequences	Annealing temp. ( C)	Length of base pairs
GAPDH Sense, 5′-CAAGGTCATCCATGACAACTTTG-3′ Antisense, 5′-GTCCACCACCCTGTTGCTGTAG-3′	61	496
IL-10 Sense: 5′-AGAGACTTGCTCTTGCACTACCAA-3′ Antisense:5′-GTAAGAGCAGGCAGCATAGCAGT-3′	60	131
IL-12p40 Sense, 5′-CAGAAGCTAACCATCTCCTGGTTTG-3′ Antisense, 5′-TCCGGAGTAATTTGGTG CTTCACAC-3′	64	251
IFN-γ Sense: 5′-GTCAACAACCCACAGGTCCAG-3′ Antisense: 5′-TTGGGACAATCTCTTCCCCA-3′	61	284

Table 2. Assessment of the stimulatory effect of ethanolic extract, water extract and berberine standard on splenocytes' proliferation.

	Splenocytes (relative absorbance of treated cells with control sample) (%)
Concentration µg/mL	Berberine standard	Water extract	Ethanolic extract
12.5	104.47 ± 2.90^a	105.82 ± 2.15^a	111.43 ± 2.74^a
25	122.85 ± 3.47^b	111.22 ± 0.81^b	116.25 ± 1.34^b
50	143.74 ± 3.75^c	140.06 ± 2.78^c	144.31 ± 2.76^c
100	169.08 ± 3.95^d	163.08 ± 3.37^d	165.78 ± 2.60^d

Note: The data are represented as the mean of triplicate experiments ±SD.

Different letters indicate significant differences in mean values, at p < 0.05. If two or more means have the same superscript letter, it means that no significant difference was detected. Mean with letter (^a) is lower than means with letter (^b), which is lower than means with letter (^c). Mean with letter (^d) has the highest value.

Table 3. Concentration of INF-γ and IL-12 in the splenocytes' culture medium with 100 µg/mL of each extract and berberine standard.

	Concentration of INF-γ and IL-12 in different extracts (pg/mL)
Assessed proteins	Control	Berberine standard	Water extract	Ethanolic extract
IFN-γ	15 ± 2.35^a	55.55 ± 3.87^c	31.11 ± 1.92^b	70 ± 2.01^dc
IL-12	10.34 ± 1.3 ^a	10.87 ± 6.5 ^a	12.53 ± 2.9 ^a	16.10 ± 1.2 ^b

Note: The data are represented as mean of triplicate experiments ±SD.

Different letters indicate significant differences in mean values, at p < 0.05. If two means or more have the same superscript letter that means no significant difference was detected. Mean with letter (^a) is lower than means with letter (^b), which is lower than means with letter (^c). Mean with letter (^d) has the highest value.

Table 4. Relative gene expression of splenocytes' IL-10, IFN-γ and IL-12 after stimulation with 100 µg/mL of each extract and berberine standard.

	Relative gene expression
Extracts	IL-10	IL-12	IFN-γ
Berberine standard	0.41 ± 0.15^a	0.19 ± 0.06^a	0.32 ± 0.14^a
Water extract	0.42 ± 0.13^a	0.32 ± 0.08^b	0.35 ± 0.14^a
Ethanolic extract	0.40 ± 0.18^a	0.35 ± 0.02^b	0.43 ± 0.17^b

Note: The data are represented as mean of triplicate experiments ±SD.

Different letters indicate significant differences in mean values, at p < 0.05. If two means or more have the same superscript letter that means there is no significant difference was detected. Mean with letter (^a) is lower than means with letter (^b), which is lower than means with letter (^c). Mean with letter (^d) is the highest value.

Table 5. Effect of B. vulgaris extract on splenocytes' parameters.

Splenocytes' parameters	Splenocytes	Stimulated splenocytes	Percentage increase
IL-12 (pg/mL)	60 ± 6	63 ± 6.1	5
IFN-γ (pg/mL)	47 ± 1.3	53 ± 1.2*↑	12.76
MHC II (%)	0.22 ± 0.021	1.21 ± 0.11*↑	450
CD11c (%)	0.09 ± 0.009	0.32 ± 0.03*↑	255.55
CD3 (%)	0.34 ± 0.035	2.02 ± 0.2*↑	494.11
LDH cell (μU/mL)	8 ± 0.8	13 ± 1.4*↑	62.5
LDH supernatant (μU/mL)	93 ± 9	70 ± 6.9*↓	−24.73
Proliferation	0.135 ± 0.011	0.16 ± 0.01*↑	18.51
NO (µmol/L)	16.3 ± 0.5	15.5 ± 0.45*↓	−4.90
IFN-γ/GAPDH	0.797 ± 0.07	0.742 ± 0.069	−6.90
IL-12/GAPDH	0.315 ± 0.029	0.397 ± 0.029*↑	26.03
IL-10/GAPDH	0.582 ± 0.055	0.523 ± 0.05	−10.13

Note: The data are represented as mean of triplicate experiments ± SD; means with asterisks are significantly different from the other group.

Significant increase (↑); significant decrease (↓).