Objective To observe the effect of electroacupuncture(EA) and moxibustion of"Dazhui"(GV 14) on the proliferation levels of the splenetic CD4

+

CD25+ CD25

+

regulatory T cells(Tregs) of H_(22) tumor-bearing mice in vitro.Methods Forty eight Balb/c mice were randomized into control

model

moxibustion and EA groups

with 12 cases in each.H_(22) tumor-bearing model was set up by hypodermic injection of H_(22) tumor cells(0.2 ml

1×10+ regulatory T cells(Tregs) of H_(22) tumor-bearing mice in vitro.Methods Forty eight Balb/c mice were randomized into control

model

moxibustion and EA groups

with 12 cases in each.H_(22) tumor-bearing model was set up by hypodermic injection of H_(22) tumor cells(0.2 ml

1×10

7

cells/ml).EA(2 Hz

2 mA) was applied to"Dazhui"(GV 14) and left"Huantiao"(GB 30) for 20 min

and moxibustion was applied to"Dazhui"(GV 14) 2 moxa-cones every time. The treatment was given from the 27 cells/ml).EA(2 Hz

2 mA) was applied to"Dazhui"(GV 14) and left"Huantiao"(GB 30) for 20 min

and moxibustion was applied to"Dazhui"(GV 14) 2 moxa-cones every time. The treatment was given from the 2

(nd)

day on after innoculation of tumor cells

once every other day

6 times altogether.After the treatment

the mice were killed by peeling off the eyeball and blood samples were collected to be separated into serum.Then

Tregs of the spleen tissues of Balb/c mice in different groups were isolated by using megnetic activated cell sorting(MACS) system to be cultured independently

and co-cultured with EA-treated serum and moxibustion-treated serum separately in culture fluid for 96 h

added with (nd) day on after innoculation of tumor cells

once every other day

6 times altogether.After the treatment

the mice were killed by peeling off the eyeball and blood samples were collected to be separated into serum.Then

Tregs of the spleen tissues of Balb/c mice in different groups were isolated by using megnetic activated cell sorting(MACS) system to be cultured independently

and co-cultured with EA-treated serum and moxibustion-treated serum separately in culture fluid for 96 h

added with

3

H-tritiate thymidine(TdR) in the culture-fluid 12 h before the end of culture

followed by collecting the cells and detecting their proliferation levels(count per minute

cpm) by using a lipid scintillation device.Results The proliferation level of Tregs in model group was elevated significantly compared to normal control group(P

<

0.05)

while in comparison with model group

those of Tregs of EA and moxibustion groups decreased considerably(P

<

0.01).After separate application of the diluted acupuncture-treated serum and moxibustion-treated serum at 1:1 and 1:8(not 1:16 and 1:32) to the cultured Tregs

their proliferation levels(cpm) in EA and moxibustion groups were obviously upregulated in comparison to those of normal control group(P

<

0.05)

and the cpm in EA group was significantly higher than that in model group(P

<

0.05)

suggesting a different action mechanism between acupuncture-moxibustion treatment and serum stimulation.Conclusion EA of"Dazhui"(GV 14) and"Huantiao"(GB 30) and moxibustion of"Dazhui"(GV 14) can effectively downregulate the proliferation level of the cultured splenetic Tregs of the tumor bearing mice.EA-treated serum and moxibustion-treated serum diluted at 1:1 and 1:8 can evidently upregulate the proliferation level of Tregs in vitro.