Chlamydia pneumoniae infections augment atherosclerotic lesion formation: a role for serum amyloid P.

Multiple reports have demonstrated an association between Chlamydia pneumoniae (Cpn) and cardiovascular disease. In this study we evaluated the effect of Cpn infections on early lesion progression in C57BL/6J mice. Since plaque formation in these mice does not develop past the initial stage, we thought these mice might be a better model for unravelling the effect of Cpn infection on early lesion type progression. C57BL/6J mice were fed an atherogenic diet and injected 10 times with 5 x 10(7) IFU Cpn or mock. At sacrifice, lesion number, size and type were analysed. To study the role of Cpn in inflammation, serum amyloid P (SAP) in plasma was determined as well as T-cells, macrophages and SAP in the lesions. In the aortic sinus of both groups, type 2 lesions were found. Cpn infection resulted in a 2.2-fold increase in total lesion size (Cpn: 10821+/-2429 microm(2)vs mock: 5022+/-1348 microm(2); p=0.04). No difference in lesion number was observed. Also, Cpn infection increased SAP in the lesions from 1.10(-4)+/-0.1.10(-4) SAP-positive cells/lesion area to 10.10(-4)+/-1.10(-4) SAP-positive cells/lesion area (p=0.05). The influx of T-lymphocytes and macrophages in the lesions as well as SAP plasma levels were not different between groups. Multiple Cpn infections resulted in a significant increase in total lesion size of C57BL/6J mice. Increase in total SAP-positive area in infected mice suggests a role for this acute-phase protein in lesion enlargement.

Secondary prevention of atherosclerosis through chlamydia pneumoniae eradication (SPACE Trial): a randomised clinical trial in patients with peripheral arterial disease.

BACKGROUND: Sero-epidemiological and experimental studies suggest that Chlamydia pneumoniae infections play an important role in the development of atherosclerosis. Clinical trials have shown contradictory results regarding the efficacy of antibiotics to prevent atherosclerosis-related complications in patients with coronary artery disease. Our aim was to study the effect of a short course of azithromycin on the incidence of cardiovascular events and peripheral vascular function in patients with stable peripheral arterial disease (PAD). PATIENTS AND METHODS: Five hundred and nine PAD-patients were randomised to receive either a 3-day course of azithromycin (500 mg daily) or placebo, with 2 years of follow-up. C. pneumoniae serology was determined at baseline. Clinical endpoints were death, coronary events (myocardial infarction, unstable angina, and/or coronary revascularization procedures), cerebral events (stroke, TIA, and/or carotid endarterectomy) and peripheral arterial complications (increased PAD-symptoms with decreased ankle-brachial index (ABPI, 0.1-point decrease after 12 months), and/or peripheral revascularization procedures). RESULTS: Five hundred and nine patients (160 women) with an atherosclerotic risk factor profile were randomised, 257 patients to azithromycin and 252 to placebo. Four hundred and forty nine patients (88%) had intermittent claudication and 60 (12%) had critical limb ischemia. By 24-month follow up, 182 patients (36%) developed 252 complications (45 deaths, 34 coronary events, 34 cerebral events and 139 peripheral arterial complications). C. pneumoniae IgA-titres were associated with the development of cardiovascular events. Nevertheless, the number of complications (131 in the azithromycin group vs. 121 in the placebo group) and the number of patients that developed complications (98 (38%) in the azithromycin vs. 84 (33%) in the placebo group) was comparable in both treatment groups. Life table analysis showed no effect of azithromycin on survival or ABPI. CONCLUSION: A short-term course of azithromycin offers no benefits for survival or ankle pressure in PAD-patients.

Chlamydia pneumoniae infection induces an unstable atherosclerotic plaque phenotype in LDL-receptor, ApoE double knockout mice.

OBJECTIVES: To study whether Chlamydia pneumoniae (Cpn) infection affects atherosclerotic plaque morphology in atherogenic (LDLr/ApoE(-/-)) mice. METHODS: In mice sacrificed 20 or 40 weeks after Cpn infection aortic arch sections were analysed for lesion and fibrous cap area and the presence of matrix metalloproteinases (MMP)-2 and -9. RESULTS: All infected mice seroconverted, demonstrated Cpn DNA in their aortas on PCR and developed atherosclerotic plaques. Infection was not associated with changes in lesion area or type, but was associated with reduced the fibrous cap area and increased MMP-2 and -9 immunoreactivity. CONCLUSION: These findings suggest that Cpn infection may predispose to plaque instability.

Screening for asymptomatic internal carotid artery stenosis and aneurysm of the abdominal aorta: comparing the yield between patients with manifest atherosclerosis and patients with risk factors for atherosclerosis only.

OBJECTIVE: The purpose of this study was to investigate whether screening for internal carotid artery stenosis (ICAS) and aneurysm of the abdominal aorta (AAA) is indicated in patients with either manifest atherosclerotic disease or with only risk factors for atherosclerosis. STUDY DESIGN: Data were obtained for 2274 patients enrolled in the SMART study, an ongoing single-center, prospective cohort study of patients referred to our vascular center with manifest atherosclerotic disease (peripheral atherosclerotic disease [PAD]; transient ischemic attack [TIA], stroke, or ICAS; AAA; angina pectoris; or myocardial infarction [MI]) or with only risk factors for atherosclerosis (diabetes mellitus, hypertension, hyperlipidemia). The presence of ICAS or AAA was determined with duplex scanning and ultrasonography. RESULTS: The prevalence of ICAS 70% or greater is low in patients with risk factors for atherosclerosis only (1.8%-2.3%), intermediate in patients with angina pectoris or MI (3.1%), and highest in patients with PAD (12.5%) or AAA (8.8%). The prevalence of AAA 3 cm or larger is low in patients with risk factors for atherosclerosis only (0.4-1.6%), intermediate in patients with angina pectoris or MI (2.6%), and highest in patients with PAD (6.5%) or TIA, stroke, or ICAS (6.5%). The prevalence of AAA larger than 5 cm is low in all of the considered patient groups. The yield of screening can be optimized through selection on the basis of simple patient characteristics. In patients with PAD, selecting those with advanced age (>54 years) increased the prevalence of ICAS to 21.8%. Selecting patients with lower diastolic blood pressure (<83 mm Hg) increased the prevalence of ICAS to 17.9%. In patients with both advanced age and lower diastolic blood pressure, the prevalence of ICAS increased to 34.7%. Selecting patients with advanced age increased the prevalence of AAA 3 cm or larger to 9.6%. In patients with TIA, stroke, or ICAS, selecting those with advanced age increased the prevalence of AAA 3 cm or larger to 8.2%. Selecting patients with taller stature (>169 cm) increased the prevalence of AAA 3 cm or larger to 9.3%. In patients with advanced age and taller stature, the prevalence of AAA 3 cm or larger increased to 13.1%. CONCLUSIONS: Screening for ICAS should be limited to patients referred with PAD or AAA, especially those with advanced age or with low diastolic blood pressure. Screening for AAA should be limited to patients referred with PAD or with TIA, stroke, or ICAS, particularly those with advanced age or tall stature. In patients referred with angina pectoris or MI and those referred with only risk factors for atherosclerosis, screening cannot be endorsed.

Chlamydophila pneumoniae (Chlamydia pneumoniae) accelerates the formation of complex atherosclerotic lesions in Apo E3-Leiden mice.

OBJECTIVE: Atherosclerosis is an inflammatory process and is characterised by the presence of T-lymphocytes in the lesions. To study the role of Chlamydophila pneumoniae (C. pneumoniae) in this process and the effect of infection on T-cell influx, we infected Apo E3-Leiden mice with C. pneumoniae and investigated the effect on lesion development and T-cell influx in atherosclerotic lesions at different time points post infection (pi). METHODS: Nine week old mice, fed an atherogenic diet, were either mock-infected or infected with C. pneumoniae and sacrificed at 1, 6 and 9 months pi. Longitudinal sections of the aortic arches of the mice were stained with hematoxylin-eosin for atherosclerotic lesion type and lesion area analysis, or with rabbit-anti-CD3(+) to detect the presence of T-cells in the atherosclerotic lesions. T-cell influx was expressed as number of T-lymphocytes/lesion area. RESULTS: At 1 month pi, type 1, 2 and 3 lesions were present. At other time points pi, more complex lesion types 4, 5a and 5b were also present. Although infection did not influence the total lesion number or area, we observed an effect of C. pneumoniae infection on lesion type. Infection resulted in a significant shift in lesion formation from type 3 to type 4 (P=0.022) at 6 months pi, and from type 4 to type 5a (P=0.002) at 9 months pi. T-cells were observed at every time point pi. At 1 month pi, a significant increase in T-cell influx in the C. pneumoniae-infected atherosclerotic lesions was observed (P=0.0005). CONCLUSION: This study shows that C. pneumoniae infection enhances the inflammatory process by increasing T-lymphocytes in the plaque and accelerates the formation of complex lesions.

Microcirculatory effects of experimental acute limb ischaemia-reperfusion.

BACKGROUND: The object of this study was to develop an animal model in which changes in microvascular haemodynamics and leucocyte-vessel wall interactions due to acute limb ischaemia-reperfusion (I/R) can be measured in the skin. Furthermore, it was investigated whether these changes are related to local muscle injury. METHODS: Male Lewis rats were subjected to unilateral limb ischaemia for 1 h (n = 8) or 2 h (n = 8) by cuff inflation, or to a sham protocol (n = 6). Intravital video microscopic measurements of leucocyte-vessel wall interactions, venular diameter, red blood cell velocity and reduced velocity (which is proportional to wall shear rate) were performed in skin venules before ischaemia and at 0.5, 1, 2, 3 and 4 h after the start of reperfusion. Oedema and leucocyte infiltration of ischaemic/reperfused skeletal muscle were quantified histologically. RESULTS: In skin venules, both 1 and 2 h of ischaemia induced a significant increase in leucocyte rolling (six and five times baseline, respectively; P < 0.05) and adherence during reperfusion (eight and four times baseline; P < 0.05). No significant increase in muscular leucocyte infiltration was detected. After an initial hyperaemic response of 180 per cent of baseline values (P < 0.05), blood flow decreased to about 60 per cent after 4 h of reperfusion in skin venules of both experimental groups. I/R induced tibial muscle oedema, the severity of which depended on the ischaemic interval (wet to dry ratio: control, 4.0; 1 h, 4.5 (P not significant); 2 h, 5.8 (P < 0.05)). CONCLUSION: A non-invasive animal model was developed that enables investigation of the consequences of acute limb I/R.

Motor dysfunction and reflex sympathetic dystrophy. Bilateral motor denervation in an experimental model.

Reflex sympathetic dystrophy (RSD) is a neuropathic pain condition most often occurring in relation to trauma to, or surgery on, an extremity. It is characterized among other things by motor disturbances such as joint stiffness and tremor. Signs and symptoms can be induced in a rat model through chronic constriction of a sciatic nerve (CCI-model). In this study the CCI-model was used to evaluate the extent of bilateral peripheral motor nerve-fiber involvement in relation to ligature localization. In 12 Lewis rats, the common sciatic nerve was loosely ligated with four chromic catgut ligatures at the midthigh level just proximal to the right sciatic trifurcation. Acetylcholinesterase (CE) histochemistry of sciatic (distal and proximal to ligation) and corresponding contralateral nerve biopsy specimens was performed at 21 days after ligation. An additional 12 rats were sham-operated and served as controls. As compared to sham-operated controls or contralateral nonligated sciatic nerves, CE histochemistry after 21 days revealed a marked decrease of CE-positive fibers in cross-sections taken from distal and proximal sciatic nerve biopsies ipsilateral to the ligatures. In addition, as compared to sham-operated controls, there was a decrease of CE-positive fibers in cross-sections taken from contralateral nonligated sciatic nerves. The present findings indicate profound motor denervation, distal as well as proximal to the ligatures. Motor denervation also affected the contralateral nonligated sciatic nerve. The evident usefulness of the CCI-model for the study of RSD places the present results in line with the concept of central nervous system involvement in the pathophysiology of RSD.

Motor denervation induces altered muscle fibre type densities and atrophy in a rat model of neuropathic pain.

Loose ligation of a sciatic nerve in rats (chronic constriction injury; CCI) provokes sensory, autonomic, and motor disturbances like those observed in humans with partial peripheral nerve injury. So far, it is unknown whether these motor disturbances result from (mechanical) allodynia or from damage to the motor neuron. These considerations prompted us to assess, in CCI rats, the density of motor axons in both the ligated sciatic nerve and the ipsilateral femoral nerve. To this end, we determined the number of cholinesterase positive fibres. It has been demonstrated previously that muscle fibre type density may be used as a measure of motor denervation and/or hypokinesia. Therefore, the myofibrillar ATPase reaction was employed to assess fibre type density in biopsies obtained from the lateral gastrocnemius muscle (innervated by sciatic nerve) and rectus femoris muscle (innervated by femoral nerve). We observed axonal degeneration of motor fibres within the loosely ligated sciatic nerve, both at an intermediate (day 21) and at a late stage (day 90) after nerve injury. The reduction in the number of motor nerve fibres was more pronounced distal to the site of the ligatures than proximal. A (less pronounced) reduction of motor fibres was observed in the ipsilateral (non-ligated) femoral nerve. In line with these findings, we observed altered fibre type densities in muscle tissue innervated by the ligated sciatic nerve as well as the non-ligated femoral nerve indicative of motor denervation rather than hypokinesia. The findings of this study suggest that the motor disorder induced by partial nerve injury involves degeneration of motor nerve fibres not only within the primarily affected nerve but also within adjacent large peripheral nerves. This spread outside the territory of the primarily affected nerve suggests degeneration of motor neurons at the level of the central nervous system.

Effects of surgical sympathectomy on skin blood flow in a rat model of chronic limb ischemia.

The role of lumbar sympathectomy in the treatment of limb ischemia secondary to arteriosclerosis obliterans has been controversial. Increased temperature and rubor of the skin, which usually follow sympathectomy, have generally been interpreted as indicative of improved nutritive skin blood flow. However, the existence of a (nonnutritive) thermoregulatory level of skin microcirculation makes such an extrapolation questionable. We investigated the total (mainly thermoregulatory) skin blood flow (TSBF) in the hindlimb of 15 male Lewis rats by means of laser Doppler flowmetry and the nutritive skin blood flow (NSBF) by means of capillary microscopy (red blood cell velocity). Transcutaneous oximetry was used to assess skin oxygenation (SO). Measurements were performed before and 2 and 28 days after ligation of the common iliac and iliolumbar artery. Subsequently, either a surgical resection of the sympathetic chain (L2-L6) was performed or a sham operation. Measurements were repeated 2 and 28 days later. For the group of 15 rats as a whole, TSBF (p < 0.05), NSBF (p < 0.05), and SO (p < 0.05) were found to be drastically reduced at day 2 after litigation compared to preligation values. This reduction partially recovered during the following weeks. TSBF (p < 0.05) and NSBF (p < 0.05), however were still reduced at day 28 after ligation compared to preligation values, whereas the SO at this time tended to be lower (p = 0.11). In the sympathectomy group the TSBF was found to be increased at day 2 (p < 0.05) and day 28 (p < 0.05) after sympathectomy, both compared to values obtained at day 28 after ligation. Sympathectomy did not have an effect on NSFB and SO. The sham procedure had no effect on the TSBF, NSBF, or SO. These results indicate that in case of lower limb ischemia, sympathectomy improves skin blood flow at the thermoregulatory but not the nutritive level of skin microcirculation. This may be related to the fact that the thermoregulatory vessels are mainly sympathetically controlled, whereas the nutritive capillaries are mainly controlled by local (nonneural) factors.

Partial peripheral neuropathy and denervation induced adrenoceptor supersensitivity. Functional studies in an experimental model.

Sciatic nerve ligation in rats (chronic constriction injury (CCI)) induces clinical signs and symptoms that mimic human conditions of neuropathic origin, such as reflex sympathetic dystrophy (RSD). Denervation-induced supersensitivity to (circulating) catecholamines has been implicated in sympathetic dysfunction in the CCI model as well as in RSD. In the present paper we studied functional properties of sympathetic innervation in subcutaneous resistance arteries, isolated from the hind paw of rats 3 weeks after ligation. Contractile responses to electric field stimulation of adrenergic nerves and exogenously administered cumulative doses of various adrenergic agonists were studied using a myograph. As compared to the contralateral side, subcutaneous arteries from the ligated side were less responsive to electrical field stimulation. Besides, as compared to the contralateral side, subcutaneous arteries from the ligated side showed increased sensitivity to alpha 1-adrenoceptor stimulation. This study demonstrates that sympathetic dysfunction in an experimental model of neuropathic pain consists of denervation-induced supersensitivity to catecholamines rather than of an afferently-induced increase in efferent sympathetic nerve impulses.

Neurogenic inflammation in an animal model of neuropathic pain.

Loose ligation of a rat sciatic nerve (chronic constriction injury (CCI) model) provokes signs and symptoms like those observed in reflex sympathetic dystrophy (RSD) patients. Primary afferent nociceptive C-fibers seem to be involved in an afferent orthodromic as well as in an efferent antidromic manner. In this study we hypothesize that consequent to development of antidromic impulses in C-nociceptive afferents, neuropeptides released from peripheral endings of these fibers, increase skin blood flow (SBF), vascular permeability, and tissue accumulation of polymorphonuclear leukocytes (PMNs). Collectively, these phenomena have been referred to as neurogenic inflammation. To investigate the presence of neurogenic inflammation in the CCI-model, we assessed skin blood flow (SBF) as well as the level of edema and accumulation of PMNs in muscle tissue obtained from the affected hindpaw. SBF was measured, by means of laser Doppler flowmetry, before ligation as well as at day 4 after ligation. At day 4, SBF measurements were performed before and after abolition of the capability of C-fibers to mediate a vasodilator response. To this end, capsaicin was applied perineurally. Increased vascular permeability was inferred from the level of edema of muscle tissue as determined by assessment of wet/dry weight ratios of muscle biopsies. PMN accumulation was investigated by enzymatic detection of myeloperoxidase (MPO) activity in muscle biopsies. Compared with preligation values, at day 4 SBF was increased more than twofold (p < 0.05). The latter response was annihilated by capsaicin application. Compared with sham operated controls, wet/dry ratios were higher in the ligated animals (1.104 vs. 1.068; p < 0.05). Likewise, when compared with sham operated controls, MPO activity was found to be increased in the ligated hindpaw (Optic Density 0.15 vs. 0.89; p < 0.001). In conclusion, the findings of this study indicate that loose ligation of a sciatic nerve induces an inflammatory response in the ipsilateral hindpaw, which most likely is mediated by release of neuropeptides from the peripheral endings of antidromically acting nociceptive C-fibres.

Reflex sympathetic dystrophy: facts and hypotheses.

Reflex sympathetic dystrophy (RSD) syndrome has been recognized clinically for many years. It is most often initiated by trauma to a nerve, neural plexus, or soft tissue. Diagnostic criteria are the presence of regional pain and other sensory changes following a noxious event. The pain is associated with changes in skin colour, skin temperature, abnormal sweating, oedema, and sometimes motor abnormalities. The clinical course is commonly divided into three stages: first (acute or hyperaemic), second (dystrophic or ischaemic), and third (atrophic) stage. The diagnosis is primarily clinical, but roentgenography, scintigraphy, thermography, electromyography and assessment of nerve conduction velocity can help to confirm the diagnosis. Although a wide variety of treatments have been recommended, the only therapies found to be effective in large studies aim at interfering with the activity of the sympathetic nervous system. To this end, efferent sympathetic nerve activity can be interrupted surgically or chemically. Alternatively, adrenoceptor blockers may be used to relieve pain. Numerous theories have been proposed to explain the pathophysiology. Sympathetic dysfunction, which often has been purported to play a pivotal role in RSD, has been suggested to consist of an increased rate of efferent sympathetic nerve impulses towards the involved extremity induced by increased afferent activity. However, the results of several experimental studies suggest that sympathetic dysfunction consists of supersensitivity to catecholamines induced by (partial) autonomic denervation. Besides, it has been suggested that excitation of sensory nerve fibres at axonal level causes release of neuropeptides at the peripheral endings of these fibres. These neuropeptides may induce vasodilation, increase vascular permeability, and excite surrounding sensory nerve fibres -- a phenomenon referred to as neurogenic inflammation. At the level of the central nervous system, it has been suggested that the increased input from peripheral nociceptors alters the central processing mechanisms.

Neurogenic inflammation and reflex sympathetic dystrophy (in vivo and in vitro assessment in an experimental model).

In the chronic constriction injury (CCI) model, signs and symptoms similar to those observed in reflex sympathetic dystrophy (RSD) can be induced by loosely ligating a rat sciatic nerve. Skin microcirculatory (inflammation-like) disorders may result from release of vasoactive neuropeptides at peripheral endings of antidromically acting nociceptive nerve fibers. These antidromic mechanisms may account for vasodilation and polymorphonuclear leukocyte (PMN) accumulation in the ligated hindpaw. We assessed skin blood flow (SBF) on the ligated side, by means of laser Doppler flowmetry, before as well as at day 4 after ligation. Postligation SBF measurements were performed before and after selective (capsaicin) conduction blockade of the ligated sciatic nerve. The extent of PMN accumulation was determined by measuring myeloperoxidase (MPO) activity in muscle biopsies obtained from the ligated and contralateral nonligated side. As compared to preligation SBF values, we observed an increase at day 4. SBF returned to preligation values consequent to capsaicin application. MPO activity, when compared to the nonligated side, was higher in biopsies obtained from the ligated side. These findings indicate that in the CCI-model, antidromically acting C-nociceptor nerve fibres increase SBF at 4 days after ligation. In addition, these antidromic mechanisms may induce an inflammatory response in the ipsilateral hindpaw, mediated by release of neuropeptides from the peripheral endings of antidromically acting C-nociceptor nerve fibers. This inflammatory response may account for various signs and symptoms as observed in the CCI model and may mirror pathophysiological mechanisms of RSD.

Systemic small-vessel disease is not exclusively related to lacunar stroke. A pilot study.

BACKGROUND AND PURPOSE: Lacunar infarcts usually results from a vasculopathy of the small vessels of the brain. It is not known whether this small-vessel disease is exclusively related to the brain or part of a more systemic small-vessel disease. In this study, patients with a lacunar stroke were investigated for manifestations of extracerebral small and large-vessel disease in comparison with cortical stroke patients. METHODS: Twenty-nine patients with a lacunar stroke, presumably caused by small-vessel disease, and 30 patients with a cortical stroke, presumably caused by large-vessel disease, entered the study. Extracerebral large-vessel disease was investigated using carotid and renal duplex scanning and Doppler sonography of the large leg vessels. Extracerebral small-vessel disease was studied from photographs of the retina, renal perfusion scintigraphy before and after angiotensin-converting enzyme inhibition, plasma renin measurement, and capillary microscopy of the nailfold. RESULTS: Vascular risk factor profile was similar in both stroke subgroups. Carotid large-vessel disease (stenosis > or =50%) was significantly less frequent among lacunar stroke patients (lacunar 3% v cortical 50%, (c)OR=0.04; 95% CI, 0.01 to 0.21, P<.01). Large-vessel disease of the renal artery (lacunar 23% v cortical 27%), and the legs (lacunar 38% v cortical 37%) was similar in both stroke groups. There was a high frequency of mild retinal arteriolosclerosis in both groups (lacunar 92% v cortical 80%). Renal blood flow changes were abnormal in 40% of the lacunar and 38% of the cortical stroke patients as a sign of renal small-vessel disease. Plasma renin concentrations did not differ between both groups. Both lacunar and cortical stroke patients had normal nailford capillary morphology, but red blood cell dynamics were reduced in both stroke groups, indicating small-vessel dysfunction. CONCLUSION: Lacunar and cortical stroke patients have both manifestations of systemic small-and large-vessel disease. Therefore, systemic small-vessel disease is not exclusively related to lacunar stroke patients who presumably have cerebral small vessel disease. A similar conclusion can be reached in cortical stroke patients.

Assessment of ischaemia of the distal part of the gracilis muscle during transposition for anal dynamic graciloplasty.

BACKGROUND: Dynamic graciloplasty is used to create a neosphincter in patients with intractable faecal incontinence. When mobilizing the distal gracilis muscle from the upper leg, the minor vascular pedicles have to be ligated. This can interfere with the vascular supply in this part of the muscle. METHODS: The arterial anatomy within the muscle was visualized by means of angiography of 11 postmortem specimens. To quantify potential acute ischaemia, blood flow in the distal gracilis muscle was measured in ten patients with laser Doppler flowmetry during mobilization of the muscle. RESULTS: Angiography showed that the main vascular pedicle and all minor pedicles drain into one and the same arterial system. After clamping of the minor vascular pedicles, blood flow (mean 25.8 (range 6.5-74.3) perfusion units) did not differ from values obtained before clamping (mean 25.4 (range 7.5-68.7) perfusion units). After a mean of 1.8 years, all muscles were vital. No correlation existed between the change in muscle blood flow and either squeeze pressure (r = -0.2) or functional outcome (r = 0.31). CONCLUSION: This study provides direct anatomical and physiological evidence of one arterial system within the gracilis muscle. It is therefore questionable whether ligation of the minor vascular pedicles is the bottleneck in human dynamic graciloplasty. An additional operation for vascular delay may be redundant. A prospective randomized clinical study should be performed to compare the functional outcome in patients with and without a delay procedure.

Skin blood flow abnormalities in a rat model of neuropathic pain: result of decreased sympathetic vasoconstrictor outflow?

Loose ligation of a sciatic nerve in rats provokes signs and symptoms like those observed in human conditions of neuropathic pain. Some of these have been associated with sympathetic dysfunction. Since the skin microcirculation in the rat is strongly influenced by sympathetic tone, abnormalities in skin blood flow may be used as an indirect measure of sympathetic dysfunction. We measured, by means of laser Doppler flowmetry, skin blood flow at the plantar surface of the rat hind paw before and after ipsilateral loose sciatic nerve ligation. We assessed basal skin blood flow as well as the vasoconstrictor response which follows cooling of the rat abdomen. The effectiveness of this response may be used as a measure of sympathetic vasoconstrictor outflow. As compared to the values obtained before ligation (= 100%): (1) the vasoconstrictor response was impaired (65%, P < 0.01) from day 1 onwards, whereas (2) basal skin blood flow was increased (171%; P < 0.01) from day 3 until day 5, and decreased (51%, P < 0.0001) from day 7 until day 28. At day 28, blockade of impulse propagation in the loosely ligated sciatic nerve (by means of lidocaine) did not increase the lowered level of skin blood flow. These findings suggest that in the chronic construction injury model loose ligation of a sciatic nerve reduces sympathetic vasoconstrictor outflow, which, in turn may induce supersensitivity of skin microvessels to catecholamines.

Effects of different durations of total warm ischemia of the gut on rat mesenteric microcirculation.

BACKGROUND: Gut injury due to ischemia and reperfusion (I/R) plays a pivotal role in many clinical conditions, such as small bowel transplantation, heart or aortic surgery in adults, and necrotizing enterocolitis in neonates. The influence of ischemic events on microcirculatory mechanisms is not well understood. Therefore, we studied, in vivo, local perfusion and leukocyte-vessel wall interactions before and after different periods of total warm ischemia of the whole gut and subsequent reperfusion in mesenteric microvessels. MATERIALS AND METHODS: Groups of pentobarbital-anaesthetized Lewis rats were subjected to 15 (n = 9), 30 (n = 12), or 60 min (n = 5) of total warm gut ischemia and 2 h reperfusion. As control a sham group (n = 10) was included. After ligating the inferior mesenteric artery, total warm ischemia was induced by clamping the superior mesenteric artery. Before and at different time periods after start of reperfusion intravital video microscopic measurements were performed. RESULTS: Rats subjected to 60 min ischemia died during the early reperfusion phase. Fifteen, 30, and 60 min ischemia induced in venules a significant decrease in blood flow, while diameter changes were not observed. This flow decrease was severe in the 15- and 30-min ischemia groups, dropping to 40 and 25% of control, respectively. Following 60 min ischemia blood flow did not exceed 10% of control. The total number of interacting leukocytes, a parameter which includes both leukocyte rolling and adhesion in venules, increased up to 5 or 10 times its control value following 15 or 30 min ischemia, respectively. Leukocyte-vessel wall interactions could not be studied in the 60-min ischemia group, due to the low blood flow. CONCLUSIONS: Even short periods of total warm ischemia of the whole gut induce severe attenuation of venular blood flow with an increase in leukocyte-vessel wall interactions. These changes increase with prolongation of the ischemic period. A 60-min period of total warm ischemia is fatal during the early reperfusion phase.

Influence of partial nerve injury in the rat on efferent function of sympathetic and antidromically acting sensory nerve fibers.

OBJECTIVE: To investigate how partial injury of a large peripheral nerve affects efferent (vasomotor) function of sympathetic and antidromically acting sensory nerve fibers. DESIGN: Randomized animal study. MATERIALS AND METHODS: We assessed, by laser Doppler flowmetry, skin blood flow (SBF) in the hindpaw of male Lewis rats before partial injury of the ipsilateral sciatic nerve (through loose ligation) as well as at an early stage (day 4) and at a later stage (day 21) after this procedure. This procedure has been reported to induce signs and symptoms like those observed in patients with causalgia. At the two time points after nerve injury, SBF was assessed before and after (chemical) blockade of sensory and nonsensory (sympathetic) sciatic nerve fibers. Furthermore, at day 21 we measured the density of sympathetic nerve fibers in footpad arteries. MEASUREMENTS AND MAIN RESULTS: At day 4, compared with preligation values, we observed an increase in SBF that was reduced by blockade of sensory nerve fibers. Subsequent blockade of nonsensory nerve fibers further reduced SBF. At day 21, SBF was decreased compared with preligation values. Blockade of sensory nerve fibers further reduced SBF, and subsequent blockade of nonsensory nerve fibers did so as well. The density of sympathetic nerve fibers was lower on the ligated side than on the nonligated side. CONCLUSIONS: Partial injury of the rat sciatic nerve causes an ipsilateral increase in SBF at an early stage, which is followed by a decrease at a later stage. At both stages, antidromically acting sensory and orthodromically acting nonsensory (sympathetic) nerve fibers are involved in the vasodilator response. At a later stage, however, neurogenic vasodilator mechanisms are overruled by a nonneurogenic vasoconstrictor mechanism. The latter may consist of supersensitivity of skin microvessels to catecholamines consequent to reduced neurogenic disposition of catecholamines.

Skin blood flow disturbances in the contralateral limb in a peripheral mononeuropathy in the rat.

Electrical excitation of nociceptive afferents in an extremity has been demonstrated to increase skin blood flow in the contralateral extremity. Hence, one would expect that loose sciatic nerve ligation, which induces an experimental painful peripheral neuropathy, may also provoke a vasodilator response in the contralateral hindpaw. On the non-ligated side, such a response may involve inhibited skin vasoconstrictor activity as well as neurogenically mediated active vasodilation. We studied skin blood flow changes in the rat hindpaw consequent to contralateral loose sciatic nerve ligation. After ligation, we also investigated whether blockade of afferent input from the ligated sciatic nerve to the spinal cord, by means of lidocaine, overrules the vasodilator response in the non-ligated paw. On the non-ligated side, we assessed the vasoconstrictor response of skin microvessels to cooling of the rat abdomen as a measure of skin vasoconstrictor activity in this paw. In order to investigate the involvement of sensory and/or non-sensory nerve fibers in the non-ligated sciatic nerve on skin blood flow abnormalities in the non-ligated paw, we studied the influence of blockade of these fibers through successive capsaicin and lidocaine application. We show that loose ligation of the sciatic nerve induces a vasodilator response in the contralateral hindpaw, which is completely abolished by blockade of afferent input from the ligated sciatic nerve. From day 1 after ligation, skin vasoconstrictor activity in the non-ligated paw was reduced, as indicated by an impaired vasoconstrictor response to cooling of the rat abdomen. Besides, blockade of sensory but not of non-sensory nerve fibers on the non-ligated side attenuated the vasodilator response in this paw. The data presented here indicate that loose ligation of the rat sciatic nerve induces a vasodilator response in the contralateral hindpaw. On the non-ligated side, this vasodilator response may involve inhibition of skin vasoconstrictor activity, as well as antidromically acting sensory nerve fibers.

Ischemia/reperfusion injury in rat mesenteric venules: red blood cell velocity and leukocyte rolling.

The authors determined the effects of 15 (n = 9) and 30 (n = 12) minutes of warm ischemia on the rat mesentery and compared the results with those of a sham-operated group (n = 10). Red blood cell velocity and number of rolling leukocytes were assessed before ischemia as well as 10, 20, 30, 60, 90, and 120 minutes after the start of reperfusion. Leukocyte rolling is considered to be an early step of the inflammatory process. Leukocytes roll along the vessel wall at a velocity that is clearly lower than that of the other blood cells. The preischemic values of red blood cell velocity and number of rolling leukocytes in the 15- and 30-minute ischemia groups did not differ from those of the sham group. In the sham group, no significant changes in red blood cell velocity and number of rolling leukocytes were observed over time. Compared with the sham group, the red blood cell velocity of the 15-minute ischemia group was significantly lower at 30, 60, 90, and 120 minutes after the start of reperfusion the number of rolling leukocytes did not differ significantly. For the 30-minute ischemia group, red blood cell velocity also was significantly lower at 20, 30, 60, 90, and 120 minutes after the start of reperfusion, and the number of rolling leukocytes was higher at 10, 20, and 30 minutes after the start of reperfusion. The results of this study indicate that short periods of total warm ischemia of the rat small bowel and subsequent reperfusion result in a significantly impaired microcirculatory blood flow in the mesentery. However, a prolonged period of ischemia is required to increase leukocyte-vessel wall interactions. In the future, this model will enable us to study the effect of pharmacological interventions during an early stage of the inflammatory response to ischemia/reperfusion in the gut.