Effect of capsulectomy on the hemodynamics and viability of random-pattern skin flaps raised on expanded skin in the pig

Skin flaps constructed on expanded skin usually include the underlying capsular tissue. It has been hypothesized that capsulectomy may jeopardize the viability of the expanded skin flap. The experiments reported herein were designed to test this hypothesis. Specifically, we studied the hemodynamics and viability of random-pattern skin flaps (8 X 20 cm) raised on delayed bipedicle flaps (group A) and on expanded skin pockets with capsulectomy at the time of flap elevation (group B) or with intact underlying capsular tissue (group C). Each group was randomly assigned to each flank in 16 pigs. Skin pockets were expanded by inflation of subcutaneous silicone tissue expanders with sterile saline (299 +/- 7 ml; X +/- SEM) over a period of 3 weeks. At the end of this period, the bipedicle flaps were constructed. Eight days later, random-pattern skin flaps were raised on bipedicle flaps and skin pockets. The length and area of skin flap viability, judged by the fluorescein dye test performed 1 day postoperatively, were not significantly different (p greater than 0.05) among groups A, B, and C (n = 31 to 32). There also were no significant differences (p greater than 0.05) in total skin capillary blood flow measured 1 day postoperatively (A = 2.6 +/- 0.4, B = 2.4 +/- 0.4, and C = 2.7 +/- 0.6 ml/min per flap; n = 15 to 16) and in skin viability assessed 7 days postoperatively (A = 74 +/- 2, B = 75 +/- 2, and C = 76 +/- 2 percent; n = 16) among delayed skin flaps and skin flaps raised on expanded skin pockets with or without capsulectomy. The results of this flap viability study were confirmed in 5 minipigs in a separate experiment. We conclude that capsulectomy did not have a detrimental effect on the hemodynamics and viability of random-pattern skin flaps raised on expanded skin. Furthermore, we hypothesize that skin flaps raised on expanded skin are similar to delayed skin flaps in that the skin blood flow is optimally augmented; therefore, the capsular tissue does not add significant blood supply to the overlying skin.     

Duration-dependent effects of nicotine exposure on growth and AKT activation in human kidney epithelial cells

Exposure to nicotine is known to cause adverse effects in many target organs including kidney. Epidemiological studies suggest that nicotine-induced kidney diseases are prevalent worldwide. However, the impact of duration of exposure on the nicotine-induced adverse effects in normal kidney cells and the underlying molecular mechanism is still unclear. Hence, the objective of this study was to evaluate both acute and long-term effects of nicotine in normal human kidney epithelial cells (HK-2). Cells were treated with 1 and 10 µM nicotine for acute and long-term duration. The result of cell viability showed that the acute exposure to 1 µM nicotine has no significant effect on growth. However, the 10 µM nicotine caused significant decrease in the growth of HK-2 cells. The long-term exposure resulted in significantly increased cell growth in both 1 and 10 µM nicotine-treated groups. Analysis of cell cycle and expression of marker genes related to proliferation and apoptosis further confirmed the effects of nicotine. Additionally, the analysis of growth signaling pathway revealed the decreased level of pAKT in cells with acute exposure whereas the increased level of pAKT in long-term nicotine-exposed cells. This suggests that nicotine, through modulating the AKT pathway, controls the duration-dependent effects on the growth of HK-2 cells. In summary, this is the first report showing long-duration exposure to nicotine causes increased proliferation of human kidney epithelial cells through activation of AKT pathway.     

Chronic oral nicotine treatment protects against striatal degeneration in MPTP-treated primates

The present studies were done to investigate the effect of long-term nicotine treatment against nigrostriatal damage in non-human primates. Monkeys were administered nicotine in drinking water for 6 months to provide chronic but intermittent delivery as with smoking. Plasma nicotine levels ranged from 10 to 15 ng/mL, which were within the range in cigarette smokers. Animals were then lesioned with low doses of the dopaminergic neurotoxin MPTP for several months while nicotine was continued. The results showed that levels of striatal tyrosine hydroxylase, dopamine transporter, vesicular monoamine transporter, dopamine and nicotinic receptors were greater in nicotine-treated MPTP-lesioned primates than in lesioned animals not receiving nicotine. Nicotine had no effect in unlesioned animals. Monoamine oxidase activity was similar in unlesioned and lesioned animals treated with or without nicotine, suggesting that nicotine did not exert its effects through changes in MPTP or dopamine metabolism. MPTP-induced cell loss in the substantia nigra was unaffected by nicotine treatment, indicating that nicotine acts at the striatal level to restore/maintain dopaminergic function. These data further support the possibility that nicotine contributes to the lower incidence of Parkinson's disease in smokers.     

Efficacy of topical nitroglycerin for random-pattern skin-flap salvage

The efficacy of topical nitroglycerin in the augmentation of random-pattern skin-flap survival was studied. Our model consisted of a standardized cranially based random skin flap on the dorsum of Sprague-Dawley rats. Nitroglycerin was delivered transdermally through a semipermeable membrane from a constant delivery system. The four study groups included preoperative and postoperative nitroglycerin, postoperative nitroglycerin, semipermeable membrane alone, and a control flap. Surviving flap areas were measured by a computer-assisted system, and groups were statistically analyzed for significance. In the rat model, treatment of a compromised random skin flap by topical nitroglycerin demonstrates no improvement in survival. In light of previous studies, this suggests a fundamental drug response difference between axial- and random-pattern skin flaps. Moreover, the use of a semipermeable membrane dressing alone showed a clear benefit (p less than 0.05) over nitroglycerin-treated and control animals.     

Effect of glucocorticoid treatment on skin capillary blood flow and viability in cutaneous and myocutaneous flaps in the pig

The effect of methylprednisolone treatment on skin-flap viability and capillary blood flow was studied in a series of four experiments. Intramuscular methylprednisolone injections (30 mg/kg per day), given in single or divided doses preoperatively or postoperatively, had no effect in augmenting skin viability in arterialized cutaneous, myocutaneous, or random skin flaps compared with the control. Capillary blood flow was studied in arterial buttock flaps and latissimus dorsi myocutaneous flaps raised on animals treated preoperatively with methylprednisolone or saline (control), and no significant difference in capillary blood flow was noted between the treatment and control flaps. It was concluded that methylprednisolone has no significant therapeutic effect either in increasing flap viability or in increasing capillary blood flow in skin flaps in pigs.     

Improved dorsal random-pattern skin flap survival in rats with a topically applied combination of nonivamide and nicoboxil

The effects of a topically applied combination of nonivamide and nicoboxil in improving skin perfusion and preventing distal flap necrosis were tested in a random-pattern dorsal skin flap model. Forty male Wistar rats were randomized into two groups (n = 20), and a standardized dorsal random-pattern skin flap was raised on each rat. Animals in the experimental group were treated with the topically applied drug combination four times per day for 6 days, whereas in the control group only a placebo ointment was applied each time. Skin flap viability was evaluated on day 7, and the extent of skin flap necrosis was compared between the two groups. The topically applied combination of nonivamide and nicoboxil resulted in a statistically significant decrease in skin flap necrosis, compared with the control group (mean percentage of skin flap necrosis in the nonivamide/nicoboxil-treated group, 22.6 +/- 6.0 percent; control group, 36.8 +/- 4.3 percent; p< 0.05). The topical combination of nonivamide and nicoboxil was effective in reducing ischemic necrosis in failing random-pattern skin flaps in this rat model. The results of this study suggest that such a topical drug application might have significant effects in the reduction of ischemic necrosis in the distal parts of skin flaps, and this treatment might also have applications as prophylactic therapy for risky skin flaps.     

Nicotine-induced vascular endothelial growth factor release via the EGFR-ERK pathway in rat vascular smooth muscle cells

Cigarette smoke has been firmly established as an independent risk factor for atherosclerosis and other vascular diseases. The proliferation and migration of vascular smooth muscle cells (VSMC) induced by growth factors have been proposed to play an important role in the progression of atherosclerosis. In the present study, we investigated the effects of nicotine, which is one of the important constituents of cigarette smoke, on vascular endothelial growth factor (VEGF) release, in rat VSMC. The stimulation of cells with nicotine resulted in a time- and concentration-dependent release of VEGF. Hexamethonium, an antagonist of nicotinic acetylcholine receptor (nAChR), inhibited nicotine-induced VEGF release. We next investigated the mechanisms by which nicotine induces VEGF release in the cells. The nicotine-induced VEGF release was inhibited by treatment with U0126, a selective inhibitor of MEK, which attenuated the nicotine-induced ERK phosphorylation. Nicotine induced a transient phosphorylation of ERK. Furthermore, AG1478, a selective inhibitor of epidermal growth factor receptor (EGFR) kinase, inhibited nicotine-induced ERK phosphorylation and VEGF release. These data suggest that nicotine releases VEGF through nAChR in VSMC. Moreover, VEGF release induced by nicotine is mediated by an EGFR-ERK pathway in VSMC. VEGF may contribute to the risk of cardiovascular diseases in cigarette smokers.     

Cigarette smoking and face lift: conservative versus wide undermining

The effects of cigarette smoking on the skin flaps of the face lift procedure are discussed. Reported elsewhere is a significant incidence of skin slough in smokers with use of wide undermining techniques. This complication is thought to be due to the vasoconstrictive effects of nicotine on the peripheral circulation. Our group has employed a conservative bilateral undermining technique in 407 face lifts. Of these, 32.4 percent were smokers and 67.6 percent were nonsmokers. No cases of skin slough were encountered. Our conservative undermining technique is briefly discussed. Among its advantages are shorter operative time, use of less local and/or general anesthesia, less intraoperative bleeding, adequate exposure for SMAS and platysmal surgery, and snugger skin closure without the risk of flap necrosis. As shown by our statistics, it is a safer procedure in smokers than the usually performed more radical procedure.     

Increased survival and vascularity of random-pattern skin flaps elevated in controlled, expanded skin

Controlled clinical tissue expansion, a new technique of providing donor tissue, results in an increase in surface area of expanded skin. The aim of the present study was to determine the effect of controlled tissue expansion on the surviving lengths of random-pattern skin flaps elevated in expanded tissue. In five pigs the surviving lengths of flaps raised in skin expanded for 5 weeks using a 250-cc rectangular Radovan-type tissue expander were compared with the survival lengths of flaps elevated in tissue in which a similar prosthesis was not expanded, bipedicle flaps delayed for 5 weeks, and control acutely raised random-pattern flaps. The expanded flaps had a mean increase in surviving length of 117 percent over control flaps, which was statistically significant. The delay flaps had an increase in survival of 73 percent over control flaps, which was also statistically significant. There was no significant difference in survival between expanded flaps and delayed flaps. Morphologic studies using radiographic techniques on one pig demonstrated increased vascularity with tissue expansion. The results of this work demonstrate that in addition to providing increased surface area with controlled expansion, flaps raised in expanded skin have a significantly augmented surviving length. The mechanism for this increased vascularity with expansion is not known at this time, but it may be due to physical forces associated with expansion acting as a stimulus for angiogenesis.     

Chronic Nicotine Treatment Reverses Hypothyroidism-Induced Impairment of L-LTP Induction Phase: Critical Role of CREB

We have previously shown that adult onset hypothyroidism impairs late-phase long-term potentiation (L-LTP) and reduces basal protein levels of cyclic-AMP response element binding protein (CREB), mutagen-activated protein kinase (MAPKp42/44), and calcium calmodulin kinase IV (CaMKIV) in area Cornu Ammonis 1 (CA1) of the hippocampus. These changes were reversed by chronic nicotine treatment. In the present study, levels of signaling molecules important for L-LTP were determined in CA1 area of the hippocampus during the induction phase. Standard multiple high-frequency stimulation (MHFS) was used to evoke L-LTP in the CA1 area of the hippocampus of hypothyroid, nicotine-treated hypothyroid, nicotine, and sham control anaesthetized adult rats. Chronic nicotine treatment reversed hypothyroidism-induced impairment of L-LTP at the induction phase. Five minutes after MHFS, Western blotting showed an increase in the levels of P-CREB, and P-MAPKp42/44 in sham-operated control, nicotine, and nicotine-treated hypothyroid animals, but not in hypothyroid animals. The protein levels of total CREB, total MAPK p42/44, BDNF, and CaMKIV were not altered in all groups 5 min after MHFS. Therefore, normalized phosphorylation of essential kinases such as P-CREB and P-MAPK p42/44 in the CA1 area of nicotine-treated hypothyroid animals plays a crucial role in nicotine-induced rescue of L-LTP induction during hypothyroidism.     

The effects of smoking on experimental skin flaps in hamsters

To study the effects of the inhalation of cigarette smoke on the survival of skin flaps, 30 Syrian Golden hamsters were divided into three groups of 10. Two of these groups were acclimatized to cigarette smoke in increasing increments for 9 weeks in standard Hamburg I smoking cages. The third group of 10 (group A) served as controls and were sham-smoked throughout the experiment. After acclimatization, one group of 10 (group B) was smoked for a further 6 weeks. A standard axial-pattern flap was then raised on the dorsum of the animals. Ten animals in group C were smoked for 6 weeks preoperatively and for 2 weeks postoperatively, at which time the animals in all groups were sacrificed. All animals survived the experiment. The flaps in control group A all survived without necrosis. Two of the 10 dorsal flaps sustained terminal necrosis in group B animals. Six of the 10 flaps resulted in significant terminal necrosis in group C animals. Statistical analysis of the results indicated a significant comparison between control group A and group C of those animals smoked throughout the experiment. We conclude from this experiment that the inhalation of cigarette smoke consistent with that of a heavy smoker (2 packs per day) has an adverse effect on wound healing of skin flaps in hamsters. Apparently, cessation of smoking even at the time of surgical preparation of the flap obviates much of the noxious effect and increase flap survival significantly.     

Reliable assessment of skin flap viability using orthogonal polarization imaging

Intraoperative evaluation of skin flap viability has primarily been dependent on clinical judgment. The purpose of this study was to determine whether an orthogonal polarization spectral imaging device could be used to accurately predict viability of random-pattern skin flaps. Orthogonal polarization spectral imaging is a newly developed technique that visualizes the microcirculation using reflected light without the use of fluorescent dyes and allows for noninvasive real-time observation of functional microvascular networks. In Sprague-Dawley rats (n = 24), three types of random skin flaps were designed with unknown zones of viability (n = 8 per group). After flap elevation, the skin flaps were evaluated by both clinical examination and orthogonal polarization spectral imaging. Areas of the flap determined to be nonviable by clinical examination were measured and marked. Orthogonal polarization spectral imaging was subsequently performed, and areas of the skin flap with stasis (i.e., cessation of red blood cell movement) in the dermal microcirculation on orthogonal polarization spectral imaging were measured and marked. The skin flaps were then secured in place. Flaps were evaluated on a daily basis for clinical signs of ischemia and necrosis. On postoperative day 7, the total amount of random skin flap necrosis was measured and recorded. Clinical examination of the random skin flaps significantly underestimated the actual amount of eventual flap necrosis, and as result was a very poor predictor of flap necrosis. By contrast, assessment of microcirculatory stasis using the orthogonal polarization spectral imaging device correlated well with the subsequent development of necrosis in all groups. In the three groups, the average amount of flap necrosis predicted by clinical examination deviated from actual necrosis by approximately 2 to 4 cm. However, the amount that orthogonal polarization spectral imaging differed from actual necrosis was 0.1 to 0.3 cm. Therefore, orthogonal polarization spectral imaging was an excellent predictor of eventual flap necrosis and much more accurate than clinical observation (p < 0.001). Intraoperative evaluation of axial and random pattern flap viability has traditionally been based on clinical examination as no other reliable, convenient test currently exists. The authors demonstrated that an orthogonal polarization spectral imaging device accurately predicts zones of necrosis in random pattern flaps by directly visualizing cessation of microcirculatory flow. Intraoperative stasis in the dermal microcirculation correlated precisely with subsequent flap necrosis. Orthogonal polarization spectral imaging was significantly more accurate than clinical examination, which consistently underestimated flap necrosis. The orthogonal polarization spectral imaging technique may have value in the intraoperative assessment of skin flap perfusion such as that required after skin-sparing mastectomy.     

The trauma of suction-assisted lipectomy cannula on flap circulation in rats.

The effect of suction-assisted lipectomy on cutaneous blood vessels of inguinal skin flaps was studied and compared in 191 rats. Different types of cannula tips were used; the number of passes was standardized. In one experiment, following suctioning, 3 X 2 cm groin island flaps based on inferior epigastric pedicles were raised and then reattached. Fluorescein dye study and microangiography were performed to evaluate flap viability. Flap survival was determined clinically and by histologic examination on the fifth postoperative day. Three-sided inguinal random-pattern flaps were raised in a second experiment and reattached following suctioning. On the fifth postoperative day, surviving flap areas were measured using standard photographs and an imaging computer and were compared with controls. Results showed that cannula passes accompanied by vacuum are harmful to vessels, while those unaccompanied by vacuum are not. The greater the number of suctioning passes, the more trauma there is to vessels and the greater is the likelihood of flap necrosis. Conical and spatula tips were more harmful to vessels than spherical, cobra, keel cobra, or Fournier tips. These results support the conclusion that suction-assisted lipectomy enhances the possibility of skin necrosis by traumatizing the vascular pedicle of a flap, especially when it is used as an adjunct to flap elevation.     

Effectiveness of a nicotine patch in helping people stop smoking: results of a randomised trial in general practice. Imperial Cancer Research Fund General Practice Research Group.

OBJECTIVE--To assess the effectiveness of 12 weeks'' treatment with a 24 hour transdermal nicotine patch in helping heavy smokers to stop smoking; also to assess the value of a specially written support booklet about smoking cessation and patch use compared with a simple advice pamphlet. DESIGN--Double blind placebo controlled randomised trial with a 2 x 2 factorial design. SETTING--19 general practices in Oxfordshire. SUBJECTS--1686 heavy smokers aged 25-64 (mean cigarette consumption 24/day; mean duration of smoking 25 years). MAIN OUTCOME MEASURE--Sustained cessation for the last four weeks of the 12 week treatment period, confirmed by saliva cotinine estimation (226/262 cases; 86.3%) or expired carbon monoxide concentration (36/262; 13.7%). Patients lost to follow up (155/1686; 9%) were assumed to have continued to smoke. RESULTS--Cessation was confirmed in 163 patients (19.4%) using the nicotine patch and 99 patients (11.7%) using the placebo patch (difference 7.6% (95% confidence interval 4.2% to 11.1%); p < 0.0001). There was no significant advantage in using the more detailed written support material. The most important adverse effect of the patch was local skin irritation, which occurred in 15.8% (133/842) and 5.1% (43/844) of patients using the nicotine and placebo patches respectively, was graded as severe in 4.8% (40) and 1.1% (nine), and was stated as a reason for withdrawal from the trial in 9.5% (80) and 2.8% (24). CONCLUSION--Nicotine patches are effective in a general practice setting with nursing support, but the extent to which this effect is sustained cannot be assessed until the results of longer term follow up are known.     

The influence of pentoxifylline on skin-flap survival

Experimental studies were performed on rats to demonstrate the effects of pentoxifylline (20 mg/kg per day), administered enterally, on skin-flap viability. This was compared to saline controls and to the alpha-blocking agent (phenoxybenzamine 1 mg/kg per day). Pentoxifylline-treated flaps had a greater than 50 percent increase in tissue survival over controls as compared to 22 percent with phenoxybenzamine. The two drugs in combination appeared to have no additional benefit over the use of pentoxifylline alone. Although a quantitative difference in red-cell flow could not be demonstrated in the experimental flaps utilizing 99mTc pertechnetate-tagged red-cell imaging, the dramatic increase in tissue viability suggests that the rheologic properties of pentoxifylline are important in altering the pathophysiology of skin-flap failure.     

On the mechanism by which antiadrenergic drugs increase survival of critical skin flaps

In order to asses the possibility that degeneration release of noradrenaline influences the survival of critical skin flaps, we studied the effect of various antiadrenergic drugs on skin-flap levels of noradrenaline, ATP, and cyclic AMP. Reserpine treatment depleted the skin flaps of noradrenaline and counteracted the fall in ATP and the cyclic AMP accumulation. Guanethidine had similar but less pronounced effects. Propranolol did not affect noradrenaline levels or depletion rate, but reduced the metabolic stimulation, as assessed by cyclic AMP levels in the flap. Phentolamine had no effect on basal noradrenaline levels, but tended to accelerate its disappearance and reduce lactate accumulation, a measure of hypoxia. All these drugs are known to increase skin-flap survival. It is suggested that they do so by, respectively, depleting the flap of its content of noradrenaline prior to operation or preventing the vasoconstriction and metabolic stimulation caused by released noradrenaline.     

Effect of radiation on skin expansion and skin flap viability in pigs

The aim of the present study was to investigate the effect of radiation treatment both on skin tissue expansion with the chronic inflation of subcutaneous expanders and on skin flap viability in surgically delayed and expanded skin in the pig. One flank in each of six pigs (initially weighing 17 +/- 1.8 kg) was randomly assigned for radiation treatment, and the contralateral flank served as a nonirradiated control. Three mirror-image, 8 x 10 cm, rectangular templates were marked on each flank; these templates were randomly assigned to the construction of a delayed skin flap (group A), a skin flap raised on expanded skin (group B), or a skin flap raised on expanded skin with a capsulectomy before flap surgery (group C). Radiation treatment was performed using sequential radiation with three fractions per week (810 cGy/fraction) for 2 weeks, with a total dose of 4,860 cGy. Twelve weeks after radiation treatment, skin expanders (8 x 10 cm) were installed subcutaneously in the locations assigned for skin expansion. Skin expansion by the inflation of subcutaneous skin expanders with saline twice weekly was started 8 weeks later and lasted for 3 weeks. Two weeks after surgical delay and the last skin expansion, 8 x 20 cm skin flaps were raised on the locations assigned for delayed skin flaps, expanded skin flaps, and expanded skin flaps with a capsulectomy. Skin flap viability was assessed 24 hours later using a fluorescein dye-staining technique. Skin expansion by the inflation of subcutaneous expanders with saline was slower (p < 0.05) in the radiated skin (39 +/- 6 ml/filling) than in the nonirradiated control skin (51 +/- 6 ml/filling). Radiation reduced the overall area of expanded skin by 23 percent (p < 0.05) compared with the control. Radiation treatment also reduced skin viability by 36 percent (p < 0.05) in the delayed skin flaps, 27 percent (p = 0.10) in the expanded skin flaps, and 36 percent (p < 0.05) in the expanded skin flaps with a capsulectomy when compared with their contralateral, nonirradiated controls. There were no significant differences in skin viability among these three types of skin flaps within the radiated and nonirradiated groups. Taken together, these observations indicate that radiation treatment reduced the effectiveness of the surgical delay procedure, the amount of subcutaneous skin expansion (by an increase in skin area), and skin flap viability. However, a capsulectomy alone did not affect the viability of skin flaps raised on expanded skin.     

Plasma neuropeptide Y levels relate cigarette smoking and smoking cessation to body weight regulation

Loss and disproportionate gain of body weight often seen respectively in smokers and quitters are believed to be due to disrupted energy homeostasis induced by nicotine, the major constituent of cigarette smoke. Energy homeostasis is suggested to be regulated by the coordinated actions of peripheral adipose tissue derived leptin and the brain hypothalamic orexigenic neuropeptide Y (NPY). While the studies probing the role of leptin and NPY in weight modulating effect of nicotine have so far been inconsistent and based largely on animal systems, there is a paucity of data involving human subjects. Here we measured the plasma levels of orexigenic neuropeptide Y (NPY) and leptin in 35 non-smokers and 31 cigarette smokers before and three months after smoking cessation. Compared to non-smokers, smokers were leaner and had reduced NPY and leptin levels. Smoking cessation resulted in a significant weight gain and increased waist circumference accompanied by increased leptin and NPY levels. NPY levels were significantly correlated with body weight (r=0.43, p<0.05), BMI (r=0.41, p<0.05), and waist circumference (r=0.37, p<0.05), while leptin correlated with BMI (r=0.42, p<0.05) and waist circumference (r=0.39, p<0.05). Association of leptin with smoking status, but not that of NPY, was lost after controlling for anthropometric parameters. Weight modulating effect of cigarette smoke may thus involve its direct action on NPY, independent of leptin. Altered leptin levels in smokers and quitters may merely reflect changes in body weight or precisely fat mass.     

Exposure to nicotine adversely affects the dendritic cell system and compromises host response to vaccination

The magnitude of Th1 cells response to vaccination is a critical factor in determining protection from clinical disease. Our previous in vitro studies suggested that exposure to the nicotine component of cigarette smoke skews the differentiation of both human and mouse dendritic cell (DC) precursors into atypical DCs (DCs differentiated ex vivo in the presence of nicotine) lacking parameters essential for the development of Th1-mediated immunity. In this study, we determined the causal relationship between nicotine-induced DC alterations and host response to vaccines. We show that animals exposed to nicotine failed to develop and maintain Ag-specific effector memory Th1 cells and Ab production to protein-based vaccine formulated with Th1 adjuvants. Accordingly, both prophylactic and therapeutic vaccines failed to protect and cure the nicotine-exposed mice from disease. More importantly, we demonstrate the nicotine-induced defects in the biological activities of in vivo DCs as an underlying mechanism. Indeed, i.v. administration of DCs differentiated in the presence of nicotine preferentially promoted the development of Ag-specific IL-4-producing effector cells in the challenged mice. In addition, DC subsets isolated from mice exposed to nicotine produced significantly less cytokines in response to Th1 adjuvants and inadequately supported the development of Ag-specific Th1 cells. Collectively, our studies suggest that nicotine-induced defects in the DC system compromises vaccine efficacy in smokers.     

Improvement of survival of skin flaps by combined gene transfer of hepatocyte growth factor and prostacyclin synthase

BACKGROUND: Increasing the local blood flow is a critical factor for long-term survival of skin flaps. Thus, a molecular therapy to increase the blood flow by means of an angiogenic factor is considered to be a useful strategy to improve skin flap survival. We focused on a combined strategy to stimulate not only angiogenesis, but also vasodilation of local microvessels, using co-transfection of the hepatocyte growth factor (HGF) and prostacyclin synthase (PGIS) genes to enhance the survival of random-pattern skin flaps. METHODS AND RESULTS: A 2 x 8 cm full thickness cranial pedicled random-pattern flap was made on the back of each 12-week-old male rat. At 3 days before operation, 400 microg of human HGF and PGIS naked plasmid DNA or control plasmid was transfected into the flaps by needle-less injection using a Shima Jet, resulting in successful expression of human HGF and PGIS in the skin flaps. Transfection of both genes into the distal half of skin flaps at 3 days prior to operation significantly increased the survival rate of skin flaps, while transfection all over the flaps did not. In addition, transfection prior to operation was more effective than simultaneous treatment. Moreover, co-transfection of these genes improved the survival area of skin flaps, accompanied by an increase in blood flow of skin flaps, even in a diabetic model. CONCLUSIONS: Overall, these results indicate that combination treatment with HGF and PGIS genes by Shima Jet could be an effective strategy to improve skin flap survival.