Lymphatic clearance of the human skin in patients with acute deep vein thrombosis using a novel fluorescent technique

The purpose of this study was to investigate lymphatic clearance of the human skin in patients with acute deep thrombosis of the femoral vein. In 13 patients with deep vein thrombosis and no other cause for swelling of the limbs, lymphatic clearance of the skin at the foot was measured. Ten microliters of fluorescein isothiocyanatedextran 150,000 were injected intradermally and the fluorescent light intensity of the deposit measured 10 min and 24 hours after injection by window densitometry. In addition, intralymphatic pressure was measured by the servo-nulling system. The results were compared with a sex- and age-matched control group. Fluorescent light intensity decreased by 23.8 +/- 12.3 arbitrary units or by a factor of 1.8 +/- 0.5 in patients with DVT after 24 hours, which was significantly less than in healthy controls (33.7 +/- 8.9 arbitrary units or by factor 5.0 +/- 4.1, p < 0.013). Intralymphatic pressure was not different between the two groups. These results indicate that lymphatic clearance is significantly reduced in the acute phase of deep venous thrombosis.     

Flow cytometric separation of gonadotrophs from dispersed rat pituitaries using a fluorescent GnRH antagonist

A gonadotropin-releasing hormone (GnRH) antagonist, [Ac-delta 3 Pro1,pFDPhe2,DTrp3,DLys6[-GnRH, was synthesized, conjugated to tetramethyl rhodamine, and found to retain GnRH antagonist activity. The fluorescent compound was used to label dispersed pituitary cells from 14-17 day-old Sprague-Dawley female rats. A subset comprising approximately 10% of the pituitary population was specifically labeled with a mean intensity of fluorescence 4.9-fold higher than the unlabeled population. The labeled cells were larger on average than the general population as inferred by forward narrow angle light scatter intensity measurements, and more granular as inferred by right angle light scatter intensity. Cells were sorted on the basis of fluorescent intensity: gonadotrophs were found to be concentrated in the rhodamine-positive fraction 5- or 6-fold relative to unfractionated cells, and 21- or 28-fold relative to rhodamine-negative fraction cells based on LH or FSH content, respectively. Gonadotrophs comprised 73 +/- 3.9% of the rhodamine-positive fraction by immunocytochemical staining. Sorted rhodamine-positive cells were cultured and found to be fully functional with respect to subsequent challenge with 30 nM GnRH. We conclude that the use of the fluorescent GnRH antagonist in conjunction with a multi-parameter cell sorter allows the purification of gonadotrophs, indicating, as expected, that these cells have a significantly higher level of GnRH binding sites than the general pituitary population. This technology should prove generally valuable in endocrine research.     

Normal pupillary size in fluorescent and bright light

STUDY OBJECTIVE: Despite its common clinical use, the range of normal pupillary size has been described only crudely. The objective of this report is to describe the distribution of normal pupillary sizes in 2 light conditions that are available in clinical settings. METHODS: Pupillary size measurements were taken from healthy patients by the principal investigator using a modified Haab scale. Measurements were obtained in areas with fluorescent lighting with an intensity of between 2,700 and 5,400 lux and by using bright handheld light sources producing a light intensity of greater than 54,000 lux. The effect of varying the type of handheld device (otoscope, ophthalmoscope, or penlight) on mean pupillary size was analyzed on the basis of intervals calculated from the t distribution. RESULTS: One hundred twenty-eight patients were enrolled, with a mean age of 35+/-9 years. The mean pupillary size in fluorescent light was 3.6+/-0.7 mm, and the mean size in bright light was 2.6+/-0.5 mm. Extreme values in fluorescent light were 2.6 mm (5th percentile) and 5.0 mm (96th percentile). Extreme values in bright light were 1.9 mm (3rd percentile) and 3.6 mm (96th percentile). The type of bright light source had no effect on pupillary size measurement. CONCLUSION: Pupillary sizes of greater than 5.0 mm or less than 2.6 mm are rare (<10%) in normal individuals in fluorescent lighting (2,700 to 5,400 lux), and sizes of greater than 3.6 mm or less than 1.9 mm are rare (<10%) in bright light.     

Effect of venous and lymphatic congestion on lymph capillary pressure of the skin in healthy volunteers and patients with lymph edema

The aim of the present study was to assess the influence of venous and lymphatic congestion on lymph capillary pressure (LCP) in the skin of the foot dorsum of healthy volunteers and of patients with lymph edema. LCP was measured at the foot dorsum of 12 patients with lymph edema and 18 healthy volunteers using the servo-nulling technique. Glass micropipettes (7-9 microm) were inserted under microscopic control into lymphatic microvessels visualized by fluorescence microlymphography before and during venous congestion. Venous and lymphatic congestion was attained by cuff compression (50 mm Hg) at the thigh level. Simultaneously, the capillary filtration rate was measured using strain gauge plethysmography. The mean LCP in patients with lymph edema increased significantly (p < 0.05) during congestion (15.7 +/- 8.8 mm Hg) compared to the control value (12.2 +/- 8.9 mm Hg). The corresponding values of LCP in healthy volunteers were 4.3 +/- 2.6 mm Hg during congestion and 2.6 +/- 2.8 mm Hg during control conditions (p < 0.01). The mean increase in LCP in patients with lymph edema was 3.4 +/- 4.1 mm Hg, and 1.7 +/- 2.0 mm Hg in healthy volunteers (NS). The maximum spread of the lymph capillary network in patients increased from 13.9 +/- 6.8 mm before congestion to 18.8 +/- 8.2 mm during thigh compression (p < 0.05). No increase could be observed in healthy subjects. In summary, venous and lymphatic congestion by cuff compression at the thigh level results in a significant increase in LCP in healthy volunteers as well as in patients with lymph edema. The increased spread of the contrast medium in the superficial microlymphatics in lymph edema patients indicates a compensatory mechanism for lymphatic drainage during congestion of the veins and lymph collectors of the leg.     

Lymphatic microangiopathy of the skin in systemic sclerosis.

METHODS: The cutaneous capillary lymphatic system in patients with systemic sclerosis was investigated using fluorescence microlymphography. The distal upper limbs of 16 healthy controls (mean age 62.3+/-13.1 yr) and 16 patients with systemic sclerosis (mean age 58.9+/-13.6 yr) were examined and the following parameters were evaluated: (a) single lymphatic capillaries; (b) lymphatic capillary network and cutaneous backflow; (c) extension of the stained lymphatics; (d) diameter of single lymphatic capillaries. RESULTS: At the finger level, lymphatic capillaries were lacking in five patients, while they were present in all controls (P < 0.05). Extension of the stained lymphatics was increased in 11 patients (8.1+/-6.0 mm) compared to the 16 healthy controls (2.0+/-1.2 mm) (P < 0.0001). Cutaneous backflow was observed in three patients (P < 0.05). At the hand level, lymphatic network extension was significantly different between patients (3.8+/-2.4 mm) and controls (1.2+/-0.8 mm) (P < 0.01); however, no significant differences were found at the forearm level. CONCLUSION: Lesional skin in patients with systemic sclerosis exhibits evidence of lymphatic microangiopathy.     

Enhanced cutaneous lymphatic network in the forearms of women with postmastectomy oedema

Postmastectomy oedema (PMO) of the arm is a common aftermath of axillary lymphatic damage during treatment for breast cancer. The aim of the present study was to quantify the forearm dermal lymphatic capillaries in order to determine whether they exhibit adaptive responses to PMO. Both forearms were examined by fluorescence microlymphography in 16 patients with oedema following treatment for breast cancer (mean swelling 25 +/- 4%) and 19 patients treated for breast cancer but without oedema. Delineated lymphatic networks were analysed stereologically. The main findings were: (1) lymphatic density at any specified distance from the injection site was greater in the swollen arm than the control arm (p < 0.01, t test); (2) taking into account the increased skin area, the total length of lymphatic capillaries in a 1-cm annulus of skin was 676 +/- 56 cm (swollen), compared with 385 +/- 30 cm (control) (p < 0.001, t test); (3) fluorescent marker was transported over a greater distance before draining deep in the swollen arm (2.74 +/- 0.33 cm) than in the control arm (1.59 +/- 0.24 cm) (p = 0.02); (4) there was no evidence of lymphatic dilatation in the swollen arm, and (5) in breast cancer patients without swelling, the arm on the side of radiotherapy/surgery (otherwise referred to as the unswollen arm) showed none of the above changes, indicating that the changes are linked to the oedema rather than being universal responses to breast cancer or its treatment. It is concluded that microlymphatic changes occur in the swollen arm, namely a local superficial rerouting of lymph drainage and either lymphangiogenesis and/or increased recruitment of dormant lymphatic vessels. Since blood capillary angiogenesis occurs in the swollen arms, and lymphangiogenesis occurs in experimental lymphoedema, there is a precedent for proposing lymphangiogenesis in PMO. An increased number of functional vessels would help to maintain the ratio of local tissue drainage capacity to filtration capacity.     

Sensitivity of Goats to a Light Pulse During the Night as Assessed by Suppression of Melatonin Concentrations in the Plasma

This study investigates the ability of a 1 h light pulse of different intensities at night to suppress plasma melatonin in goats. Six female Saanen dairy goats, about 2 yr old, were housed in a light-tight shed. The goats were habituated for 1 wk to an 8L:16D photoperiod (40.70 +/- 4.16 microW/cm2; 137 +/- 14 lux), lights on 0800 h. A 1 h light pulse, of different intensity on each occasion, was given from 1900 to 2000 h. Light intensity was measured by using a lux meter (mean of 36 measurements at goat's eye level). Five different light intensities were given during December in the order 4.22 +/- 0.62 microW/cm2 (14.2 +/- 2.1 lux), 0.68 +/- 0.09 microW/cm2 (2.3 +/- 0.3 lux), 0.26 +/- 0.004 microW/cm2 (0.87 +/- 0.14 lux), darkness, 40.70 +/- 4.16 microW/cm2 (137 +/- 14 lux), with 1-3 d between treatments. The goats were bled hourly from 1500 to 1900 h and every 15 min from 1900 to 2100 h, and a last bleed occurred at 2200 h. Dark-phase samples were taken in dim red light (less than 0.03 microW/cm2; 0.1 lux). Plasma was assayed for melatonin by radioimmunoassay. Suppression of melatonin concentrations increased as light intensity increased as follows: Darkness, 0%; 0.26 +/- 0.004 microW/cm2; 0%; 0.68 +/- 0.09 microW/cm2; 43.1%; 4.22 +/- 0.62 microW/cm2, 71.1%; 40.70 +/- 4.16 microW/cm2, 81.2%. Suppression was significant (P less than 0.05) at light intensities greater than 0.68 microW/cm2, 2.3 lux. A hyperbolic relationship existed between percent suppression and light intensities.     

Dextran sulfate sodium-induced acute colitis impairs dermal lymphatic function in mice

AIM: To investigate whether dermal lymphatic function and architecture are systemically altered in dextran sulfate sodium (DSS)-induced acute colitis.METHODS: Balb/c mice were administered 4% DSS in lieu of drinking water ad libitum for 7 d and monitored to assess disease activity including body weight, diarrhea severity, and fecal bleeding. Control mice received standard drinking water with no DSS. Changes in mesenteric lymphatics were assessed following oral administration of a fluorescently-labelled fatty acid analogue, while dermal lymphatic function and architecture was longitudinally characterized using dynamic near-infrared fluorescence (NIRF) imaging following intradermal injection of indocyanine green (ICG) at the base of the tail or to the dorsal aspect of the left paw prior to, 4, and 7 d after DSS administration. We also measured dye clearance rate after injection of Alexa680-bovine serum albumin (BSA). NIRF imaging data was analyzed to reveal lymphatic contractile activity after selecting fixed regions of interest (ROIs) of the same size in fluorescent lymphatic vessels on fluorescence images. The averaged fluorescence intensity within the ROI of each fluorescence image was plotted as a function of imaging time and the lymphatic contraction frequency was computed by assessing the number of fluorescent pulses arriving at a ROI.RESULTS: Mice treated with DSS developed acute inflammation with clinical symptoms of loss of body weight, loose feces/watery diarrhea, and fecal blood, all of which were aggravated as disease progressed to 7 d. Histological examination of colons of DSS-treated mice confirmed acute inflammation, characterized by segmental to complete loss of colonic mucosa with an associated chronic inflammatory cell infiltrate that extended into the deeper layers of the wall of the colon, compared to control mice. In situ intravital imaging revealed that mice with acute colitis showed significantly fewer fluorescent mesenteric lymphatic vessels, indicating impaired uptake of a lipid tracer within mesenteric lymphatics. Our in vivo NIRF imaging data demonstrated dilated dermal lymphatic vessels, which were confirmed by immunohistochemical staining of lymphatic vessels, and significantly reduced lymphatic contractile function in the skin of mice with DSS-induced acute colitis. Quantification of the fluorescent intensity remaining in the depot as a function of time showed that there was significantly higher Alexa680-BSA fluorescence in mice with DSS-induced acute colitis compared to pre-treatment with DSS, indicative of impaired lymphatic drainage.CONCLUSION: The lymphatics are locally and systemically altered in acute colitis, and functional NIRF imaging is useful for noninvasively monitoring systemic lymphatic changes during inflammation.     

Postural effects on interstitial fluid pressure in humans

BACKGROUND: Direct assessment of the effect of postural changes on interstitial fluid pressure (IFP) in the human skin under physiological conditions is important for the understanding of mechanisms involved in diseases resulting in lower limb edema. Previous techniques to measure IFP had limitations of being invasive, and acute measurements were not possible. Here we describe the effect of postural changes on IFP in the skin of the foot using the minimally invasive servonulling technique. RESULTS: Measurements were performed in 12 healthy subjects. IFP (means +/- SD) was significantly higher in the sitting (5.1 +/- 2.9 mm Hg) than in the supine position (-0.3 +/- 3.6 mm Hg, p = 0.04) when measured in the sitting position first. The difference between the sitting and the supine position was not significant when measurements were taken in the supine position first [from 1.0 +/- 4.3 (supine) to 3.6 +/- 6.7 mm Hg (sitting), p = 0.46]. Spontaneous low-frequency pressure fluctuations occurred in 58% of the recordings during sitting, which was almost twice as frequent as in the supine position (33%; p = 0.001), while no effects on lymphatic capillary network extension were observed (p = 0.12). CONCLUSION: Using the servonulling micropressure system, postural effects on IFP can be directly assessed. IFP is higher in the sitting position, but differences are influenced by the time in the upright position.     

Microlymphatic aneurysms in patients with lipedema.

"Lipedema," a special form of obesity syndrome, represents swelling of the legs due to an increase of subcutaneous adipose tissue. In 12 patients with lipedema of the legs and in 12 healthy subjects (controls), fluorescence microlymphography was performed to visualize the lymphatic capillary network at the dorsum of the foot, at the medial ankle, and at the thigh. Microaneurysm of a lymphatic capillary was defined as a segment exceeding at least twice the minimal individual diameter of the lymphatic vessel. In patients with lipedema, the propagation of the fluorescent dye into the superficial lymphatic network of the skin was not different from the control group (p > 0.05). In all 8 patients with lipedema of the thigh, microaneurysms were found at this site (7.9 +/- 4.7 aneurysms per depicted network) and in 10 of the 11 patients with excessive fat involvement of the lower leg, multiple microlymphatic aneurysms were found at the ankle region. Two obese patients showed lymphatic microaneurysms in the unaffected thigh and in only 4 patients were microaneurysms found at the foot. None of the healthy controls exhibited microlymphatic aneurysms at the foot and ankle, but in one control subject a single microaneurysm was detected in the thigh. Multiple microlymphatic aneurysms of lymphatic capillaries are a consistent finding in the affected skin regions of patients with lipedema. Its significance remains to be elucidated although its occurrence appears to be unique to these patients.     

Increased dermal lymphatic density in the human leg compared with the forearm.

During orthostasis, capillary filtration pressure is higher in the leg than in the arm due to the effect of gravity. We investigated the possibility that the lymphatic network in leg skin might be adapted to cope with a greater fluid load. The dermal lymphatics of the forearm and lower leg were studied in white-skinned and brown-skinned men using fluorescence video microscopy. From video print lymphangiograms the following were determined: lymphatic length density at a series of radii from the centre of the lymphangiogram (LDr); maximum lymphatic density (LDmax); total length of lymphatic vessel (LL); maximum spread of lymphatic vessel; number and size of lymphatic rings (continuous circuits of vessel); and vessel diameter. There were no differences between the two racial types, but clear differences between the arm and leg. In the leg, mean (+/- SD) peak LDr (25.13 +/- 5.65 cm-1), LDmax (32.95 +/- 6.89 cm-1), LL (40.17 +/- 27.42 cm), and spread (1.39 +/- 0.08 cm) were all significantly higher than in the arm (18.03 +/- 5.48 cm-1, 23.91 +/- 7.21 cm-1, 11.76 +/- 5.47 cm, and 1.00 +/- 0.05 cm respectively, P 相似文献   

The rat glomerular filtration barrier does not show negative charge selectivity

OBJECTIVE: To characterize the effects of size, shape, and negative charge on the transport of macromolecules across the glomerular capillary wall by using the sieving curves (fractional clearance vs. solute molecular radii) of fluorescent polydispersed polysaccharide tracers. METHODS: Glomerular fractional clearances (FC) were measured with fluorescent neutral [isoelectric point (pI) = 7.3 +/- 0.2] and negatively charged (pI = 3.5 +/- 0.4) dextrans (DEX) in comparison with negatively charged (pI = 4.8 +/- 0.3) hydroxy ethyl starch (HES) and (pI = 4.6 +/- 0.1) bovine serum albumin (BSA) in Sprague-Dawley and Fischer 344/Brown Norway rats. FCs (n = 53) were measured by using the urinary clearance of (14)C-inulin to determine the glomerular filtration rate. The relative uptake of each fluorescent probe by endothelial and renal proximal tubule epithelial (LLC-PK(1)) cells, in vitro, was measured microscopically by using a cooled (-25 degrees C) CCD camera. RESULTS: The sieving curves for randomly coiled neutral and negatively charged DEX probes were identical. These FC values were 6-fold greater than those for HES and 200-fold above similarly sized fluorescent BSA. The polysaccharide probes did not show significant binding to serum proteins. The uptake of BSA by LLC-PK(1) cells was 20- to 100-fold greater than that for neutral or negatively charged macromolecules. CONCLUSIONS: These findings indicate that the rat glomerular filtration barrier restricts the transport of polysaccharide macromolecules as a function of their size and configuration but not negative charge.     

Use of the disappearance rate for the estimation of lymphatic absorption during CAPD.

Several studies are discussed which investigated the usefulness of the disappearance rate of macromolecules from the peritoneal cavity for estimating convective fluid loss from the peritoneal cavity into the peritoneal lymphatic system. It is shown that dextrans are removed from the peritoneal cavity by a size-independent process at a mean rate of 1.37 +/- 0.15 ml/min, whereas the clearance from blood to dialysate of dextrans is size-dependent. The fluid removal rate estimated by the difference in bidirectional transport of inulin (1.79 +/- 0.38 ml/min; p < 0.0005) was of the same order of magnitude as has been found using the removal rate of macromolecules from the peritoneal cavity. Also, the role of local accumulation of macromolecules was studied during continuous administration of dextrans. No differences were found in the dextran disappearance rate before and after saturation of the peritoneal interstitium with dextran (1.1 +/- 0.6 vs. 1.0 +/- 0.4 ml/min). During a study using a hypoosmotic solution we calculated a net transcapillary backfiltration of fluid, whereas the dextran removal rate was in the same order of magnitude as found using commercially available dialysate. In our opinion, the disappearance rate of macromolecules is an estimate of convective fluid loss from the peritoneal cavity into the peritoneal lymphatic system.     

Fluorescence microlymphography: diagnostic potential in lymphedema and basis for the measurement of lymphatic pressure and flow velocity

Fluorescence microlymphography (FML) is an almost atraumatic technique used to visualize the superficial skin network of initial lymphatics through the intact skin of man. Visualization was performed with an incident light fluorescence microscope following subepidermal injection of minute amounts of FITC-dextran 150,000 using microneedles. Emanating from the bright dye depot, the surrounding network of microvessels is filled, documentation performed by photography or video film. In congenital Milroy lymphedema, a lack of microlymphatics (aplasia) is typical while in other primary lymphedemas and in secondary lymphedema after mastectomy or irradiation of proximal lymph nodes, the network remains intact but the depicted area is enlarged. Lymphatic microangiopathy characterized by obliterations of capillary meshes or mesh segments develops in phleboedema with trophic skin changes, progressive systemic sclerosis and Fabry's disease. In lipedema, lymphatic microaneurysms are stained. Microlymphatic pressure may also be measured using FML. For this purpose, glass micropipettes are inserted into the capillaries by means of a micromanipulator and pressure is determined by the servo-nulling technique. Normal subjects produced significantly lower pressure (7.9 +/- 3.4 mmHg) compared to patients with primary lymphedema (15.0 +/- 5.1 mmHg, p<0.001). This characteristic lymphatic hypertension may be improved by complex physiotherapy or local application of prostaglandins. Additionally, a modification of the FML procedure can be used to measure lymphatic capillary flow velocity in controls and patients. FML is suited to confirm the clinical diagnosis of lymphedema, contributes to distinguish among various forms of edema, and is useful in clinical research. In addition, FML has also become a tool for experimental animal studies including the depiction of gastric microlymphatics, the measurement of flow velocity in the naked mouse tail, and in evaluation of lymphangiogenesis in a model of Milroy disease.     

Reticulocyte counts in the aged

Reticulocytes were measured on an automated reticulocyte counter in five groups; healthy adults, non-elderly patient with low hematopoiesis, adults with anemia, healthy elderly persons and elderly patients with anemia. The reticulocyte percent, absolute reticulocyte count, and reticulocyte composition as classified by fluorescence intensity (highly, moderately, and slightly fluorescent cells) were calculated. The healthy adults had a reticulocyte count of 0.70 +/- 0.55%, an absolute reticulocyte count of 4.36 +/- 1.90 X 10(4)/microliters, 2.33 +/- 1.95% highly fluorescent cells, 18.73 +/- 5.07% of moderately fluorescent cells, and 78.82 +/- 6.55% of slightly fluorescent cells. Patients with low hematopoiesis had lower counts except for the percentage of slightly fluorescent cell. Aged persons without anemia showed no differences in reticulocytes from healthy adults. However, elderly patients with anemia had a low reticulocyte count; there was no tendency towards an increase in the percentage of highly fluorescent cells or a decrease in the percentage of slightly fluorescent cells. Their reticulocyte percent was not significantly higher than healthy controls, suggesting that anemia observed in the aged arises from low hematopoietic activity in the bone marrow.     

Effect of a short course of rhDNase on cough and mucociliary clearance in patients with cystic fibrosis

The aim of the study was to measure the effect of a short course of recombinant human deoxyribonuclease I (rhDNase) on ciliary and cough clearance in a group of cystic fibrosis patients, using a radioaerosol and gamma camera technique. Patients were initially randomized to receive either rhDNase (2.5 mg qd) or placebo. Following the measurement of baseline clearance, patients were given a 7-day course of either rhDNase or placebo. The patient then returned on the seventh day for follow-up clearance measurements. This was followed by a 2-week washout period before the whole process was repeated with the alternative inhalation solution. On each of the study days, mucociliary clearance was initially measured for a period of 60 min (IC). This was followed by cough clearance (CC) measurements for 30 min, during which patients were requested to cough a total of 120 times. Post-cough clearance (PCC) was then measured for a further 60 min. Thirteen patients completed the study. Patients' age ranged between 18-38 years, and they had baseline values of FEV(1) of 27-103% of predicted values. Following completion of the course of rhDNase, there was a mean percent increase from baseline of 7.5% for FEV(1) and 5.4% for FVC% (P = 0. 03). There was a small, nonsignificant increase in IC (6.2 +/- 3.6%) on the rhDNase arm compared with the placebo arm (-2.3 +/- 2.9%), P = 0.1. No changes were seen in either CC (1.0 +/- 3.2% [rhDNase] vs. 1.9 +/- 2.4% [placebo], P = 0.9) or PCC (-0.7 +/- 1.5% [rhDNase] vs. 0.9 +/- 1.7% [placebo], P = 0.3). Patients who achieved a 10% or greater improvement in FEV(1) (n = 5) in response to rhDNase did not show any greater change in clearance than nonresponders. In conclusion, we were unable to demonstrate any improvements in either ciliary or cough clearance in response to a short course of rhDNase. The mechanism of action of this drug in vivo remains uncertain.     

Pharmacokinetic approach to the estimation of hepatic removal of insulin

We simultaneously measured hepatic insulin removal invasively and estimated hepatic clearance and extraction of insulin pharmacokinetically from cardiac output and peripheral plasma concentrations (relatively) noninvasively. The invasive methods involved continuous electromagnetic measurements of portal venous and hepatic arterial blood flow and simultaneous intermittent sampling of blood from the portal and hepatic veins and femoral artery for assay of insulin concentrations. The noninvasive method assumed that hepatic plasma flow is proportional to cardiac output and that hepatic clearance is a constant fraction of total body clearance of insulin. In anesthetized dogs (n = 6), endogenous insulin was suppressed with somatostatin (800 ng/kg/min) while biosynthetic human insulin (0.25, 0.50, and 1.00 mU/kg/min) was infused to steady state during three consecutive 90-min periods. Insulin concentrations were directly proportional to the infusion rate (p less than 0.01). Hepatic blood flow accounted for 20 +/- 2% of cardiac output. Measured hepatic clearance accounted for 51 +/- 5% of total body clearance of insulin and correlated with the pharmacokinetic estimates (p less than 0.01); the estimates of hepatic clearance ranged from 91 to 114% of the measured values. We conclude that this pharmacokinetic approach, which requires only samples of peripheral blood and estimates of hepatic blood flow, may be used to study the hepatic removal of insulin relatively noninvasively.     

Effects of melatonin on the thermoregulatory responses to intermittent exercise

We examined the effects of a single 2.5-mg dose of melatonin on the thermoregulatory and circulatory responses to intermittent exercise at a room temperature of 27.2+/-0.4 degrees C (mean+/-S.D.), a relative humidity of 55+/-3% (mean+/-S.D.), and a light intensity of 200-300 lux. In a double-blind cross-over study, six male participants ingested either melatonin or placebo at 11:45 hr. Participants then rested in a semi-supine position for 75 min and completed an intermittent running protocol for 66 min at alternating intensities of 40, 60 and 80% of maximal oxygen uptake. Rectal and mean skin temperature, heart rate, blood pressure, skin blood flow, subjective alertness and sleepiness, ratings of perceived exertion (RPE) and thermal strain were recorded. No effects of melatonin were found on these variables measured during the resting period (P>0.10). During exercise, melatonin was found to moderate the increase in rectal temperature by approximately 0.25 degrees C (P=0.050) and magnify the increase in skin blood flow (P=0.047). Postexercise systolic blood pressure was 7.8+/-2.5 mmHg (mean+/-S.D.) lower than before the exercise in the melatonin trial; a change which differed significantly to that in the placebo trial (P=0.018). Melatonin did not influence subjective alertness and sleepiness before or after exercise and did not change the responses of mean skin temperature, RPE and thermal strain during the exercise (P>0.10). In summary it is apparent that a 2.5-mg dose of melatonin has hypothermic, but not soporific, effects during 66 min of intermittent exercise performed under moderate heat stress. Whether such effects improve endurance athletic performance in hot conditions remains to be confirmed. Our data also suggest that postexercise systolic hypotension is more marked after ingestion of melatonin.     

Documentation of pulmonary capillary permeability in the adult respiratory distress syndrome accompanying human sepsis.

Current evidence suggests that pulmonary edema accompanying human sepsis may result either from changes in the serum oncotic and hydrostatic pressures or an increase in the permeability of the pulmonary microvasculature. In this study, we compared the "clearance" of injected 131I-labeled human serum albumin from blood to bronchoalveolar secretions in intubated patients with pulmonary edema secondary to sepsis or myocardial infarction. A significantly increased mean +/- SE clearance of the radionuclide was seen in patients with sepsis (0.34 +/- 0.03 ml per hour) compared to those with myocardial infarction (0.043 +/- 0.008 ml per hour) (P less than 0.001), although both groups had similar degrees of edema on chest radiographs. Because the patients with sepsis had no severe decrease in serum oncotic pressure (18.4 +/- 5.0 mm Hg) or evidence of left heart failure, as determined by the pulmonary wedge pressure (11.0 +/- 6.8 mm Hg), we concluded that the genesis of the pulmonary edema in sepsis was due to an increase in pulmonary microvascular permeability, as measured by the increased clearance of 131I-labeled human serum albumin.     

Perimuscular connective tissue contains more and larger lymphatic vessels than the shallower layers in human gallbladders

AIM: To clarify whether perimuscular connective tissuecontains more lymphatic vessels than the shallowerlayers in human gallbladders.METHODS: Lymphatic vessels were stainedimmunohistochemically with monoclonal antibody D2-40,which is a specific marker of lymphatic endothelium, inrepresentative sections of 12 normal human gallbladdersobtained at the time of resection for colorectal carcinomaliver metastases. In individual gallbladder specimens,nine high-power (× 200) fields with the highestlymphatic vessel density (LVD), termed "hot spots", wereidentified for each layer (mucosa, muscle layer, andperimuscular connective tissue). In individual hot spots,the LVD and relative lymphatic vessel area (LVA) weremeasured microscopically using a computer-aided imageanalysis system. The mean LVD and LVA values for thenine hot spots in each layer were used for statisticalanalyses.RESULTS: In the mucosa, muscle layer, andperimuscular connective tissue, the LVD was 16.1 ± 9.2,35.4 ± 15.7, and 65.5 ± 12.2, respectively, and the LVAwas 0.4 ± 0.4, 2.1 ± 1.1, and 9.4 ± 2.6, respectively.Thus, both the LVD and LVA differed significantly (P <0.001 and P < 0.001, respectively; Kruskal-Wallis test)among the individual layers of the wall of the gallbladder,with the highest LVD and LVA values in the perimuscularconnective tissue. Most (98 of 108) of the hot spotswithin the perimuscular connective tissue were locatedwithin 500 μm of the lower border of the muscle layer.CONCLUSION: The perimuscular connective tissuecontains more and larger lymphatic vessels than the shallower layers in the human gallbladder. This observation partly explains why the incidence of lymph node metastasis is high in T2 (tumor invading the perimuscular connective tissue) or more advanced gallbladder carcinoma.