Radiographic and clinical changes of the patellar tendon after tibial plateau leveling osteotomy 94 cases (2000-2003).

Patellar tendon thickening (PTT) and patellar tendinosis (PTS) have been discussed in the veterinary literature as a post-operative complication of tibial plateau leveling osteotomy (TPLO). The purpose of this study was to define radiographic PTT, determine the frequency of and risk factors for PTT and PTS, and describe the clinical and histopathological findings of PTS after TPLO. We hypothesized that the location of the osteotomy alters forces placed on the patelloar tendon resulting in PTT or PTS. Radiographs and medical records from 83 dogs undergoing 94 TPLO procedures were retrospectively evaluated. Two months post-operatively, 19 dogs (20.2%) had a normal patellar tendon or mild PTT, 51 (54.3%) had moderate PTT, and 24 (25.5%) had severe PTT. Seven of the 24 dogs (7.4%) with severe PTT had clinical signs consistent with PTS. Only dogs with severe PTT developed PTS (p < 0.0001). The risk factors for the development of PTT include: a cranial osteotomy, a partially intact cranial cruciate ligament (CCL) in conjunction with a cranial osteotomy, and post-operative tibial tuberosity fracture. The only risk factor identified for the development of PTS was a partially intact CCL. Four dogs with PTS improved with conservative therapy and one improved with surgical treatment. Two dogs had tendon biopsies with histopothological review that showed tendon degeneration with lack of inflammation. As only the dogs with severe PTT develop PTS, a caudal osteotomy for the prevention of PTT and subsequent PTS is recommended.

Interstitial fluid pressure and blood flow in canine osteosarcoma and other tumors.

This study aims to characterize interstitial fluid pressure and blood flow in naturally occurring appendicular bone tumors in dogs because high pressure may influence the response of tumors to chemotherapy and radiation therapy. Eighteen client-owned dogs with naturally occurring appendicular bone tumors were included in this study. At the time of surgical biopsy, interstitial fluid pressure and blood flow were measured using wick-in-needle probes and laser Doppler flowmetry, respectively, within the soft tissue and bony components of the lesions and in normal muscle. Interstitial fluid pressure within the bony and soft tissue components of the tumors was significantly higher than interstitial fluid pressure in normal muscle. Blood flow in the bony component of the tumors was significantly lower than blood flow in normal muscle. There was no significant difference between blood flow in the soft tissue component of the tumors compared with that in normal muscle. Appendicular bone tumors in dogs have significantly higher interstitial fluid pressure and lower blood flow than do adjacent, unaffected soft tissues. The higher interstitial fluid pressure and lower blood flow may reduce tissue oxygenation and impede drug delivery. The effects of increased interstitial fluid pressure and decreased blood flow should be considered in the formulation of treatment strategies for the clinical management of appendicular bone tumors.

"Artificial lymphatic system": a new approach to reduce interstitial hypertension and increase blood flow, pH and pO2 in solid tumors.

A mechanical drainage system, the "artificial lymphatic system" (ALS), consisting of a vacuum source and drain, is evaluated for its ability to aspirate the interstitial fluids responsible for the elevated interstitial fluid pressure (IFP) observed in solid tumors. IFP, pH, and pO2 radial profiles were measured before and after aspiration using wick-in-needle (WIN) probes, needle pH and oxygen electrodes, respectively. Laser Doppler flowmetry measured temporal changes in blood flow rate (BFR) at the tumor surface during aspiration. The WIN probe and IFP profile data were analyzed using numerical simulation and distributed mathematical models, respectively. The model parameter, P(E), reflecting central tumor IFP, was reduced from 15.3 to 5.7 mm Hg in neuroblastoma and from 13.3 to 12.1 mm Hg in Walker 256, respectively, following aspiration. The simulation demonstrated that spatial averaging inherent in WIN measurements reduced the calculated magnitude of the model parameter changes. IFP was significantly lower (p<0.05), especially in regions surrounding the drain, and BFR was significantly higher (p<0.05) following 25 and 45 min of aspiration, respectively; pH and pO2 profiles increased following aspiration. The experimental and mathematical findings suggest that ALS aspiration may be a viable way of reducing IFP and increasing BFR, pO2, and pH and should enhance solid tumor chemo and radiation therapy.

Enhancing the uptake of chemotherapeutic drugs into tumors using an "artificial lymphatic system".

This paper presents findings from uptake studies to evaluate the ability of an "artificial lymphatic system" (ALS) to enhance large and small molecular weight drug transport into solid tumors and the therapeutic effect of the additional drug on their growth. These studies also served to test the effectiveness of an implantable multidrain ALS. Walker 256, Neuroblastoma, and Sarcoma dual-tumor models were used to evaluate the effect of ALS aspiration on the uptake of 3F8 monoclonal antibody, and doxorubicin. A tumor shrinkage experiment using Walker 256 dual tumors was used to evaluate the efficacy of an implantable ALS with cyclophosphamide chemotherapy. Drug uptake significantly increased in all aspirated tumors; 3F8 uptake was enhanced 37.4% in the Walker and 93.1% in the Neuroblastoma tumor lines (p<0.05). Doxorubicin uptake increased 23.2% in Sarcoma tumor (p<0.05). The shrinkage study demonstrated that one-drain aspirated tumors shrank 90% faster (p<0.01) than control tumors, while three-drain aspirated tumors shrank 123% faster than control tumors (p<0.01).

The timing and nature of neovascularization of jejunal free flaps: an experimental study in a large animal model.

The present study was designed (1) to determine whether a free jejunal transfer in a large animal model can develop collateral circulation that is adequate to maintain viability after division of the pedicle and (2) to determine the earliest time pedicle ligation is safe after transplantation. A 15-cm jejunal segment was transferred to the necks of 18 dogs weighing 25 to 35 kg. The bowel segment was inset longitudinally under the skin on one side of the neck, partially covered by the neck muscles, and the mesenteric vessels were anastomosed to recipient vessels in the neck. The proximal and distal bowel stomas were exteriorized through skin openings 12 cm apart and matured. The dogs were subjected to ligation of the vascular pedicle at different intervals: postoperative day 7 (group I, n = 3), day 14 (group II, n = 5), day 21 (group III, n = 5), and day 28 (group IV, n = 5). Blood perfusion was measured in the proximal and distal bowel stomas before pedicle division (control) and 24 hours later using hydrogen gas clearance and fluorescein dye. Bowel necrosis was analyzed using planimetry. The bowel was also stained with hematoxylin and eosin and factor VIII, and new blood vessels were counted. Mean values (+/- standard deviation) were compared with control values for each test and with normal values in the intact bowel using analysis of variance with Neumann-Keuls post-hoc test for multiple comparisons. No jejunal free flaps survived when the vascular pedicle was divided 1 week postoperatively. Bowel survival was 60 percent at 2 weeks, 83 percent at 3 weeks, and 100 percent at 4 weeks. Hydrogen gas clearance values (ml/min/100 g) were 49.6 +/- 8.7 in the mucosa of the intraabdominal jejunum and 37.9 +/- 9.4 in the jejunum that was transferred to the neck before division of the pedicle. Twenty-four hours after pedicle division, hydrogen gas clearance values were 2.8 +/- 6.4 in group I (p < 0.05), 22.4 +/- 12.4 in group II, 23.9 +/- 9.3 in group III, and 34.2 +/- 7.5 in group IV. FluoroScan readings in the transferred jejunum were 201 +/- 7.2 in the control group, 9.3 +/- 2.8 in group I (p < 0.05), 79.1 +/- 10.6 in group II, 66.2 +/- 7.3 in group III, and 164 +/- 11.9 in group IV. New vessel formation as identified by factor VIII staining correlated with increasing bowel perfusion and flap survival rate. Bowel neovascularization, perfusion, and survival increased progressively 1 week after transfer. Significant portions of the transferred bowel will neovascularize and survive as early as 2 weeks postoperatively. However, a minimum of 4 weeks before ligation of the pedicle is necessary to maximize flap perfusion and guarantee survival.

Imaging herpes virus thymidine kinase gene transfer and expression by positron emission tomography.

We report a series of studies that assess the feasibility and sensitivity of imaging of herpes virus type one thymidine kinase (HSV1-tk) gene transfer and expression with [124I]-5-iodo-2'-fluoro-1-beta-D-arabinofuranosyluracil ([124I]-FIAU) and positron emission tomography (PET) and the ability of [124I]-FIAU-PET imaging to discriminate different levels of HSV1-tk gene expression. Studies were performed in rats bearing multiple s.c. tumors derived from W256 rat carcinoma and RG2 rat glioma cells. In the first set, we tested the sensitivity of [124I]-FIAU-PET imaging to detect low levels of HSV1-tk gene expression after retroviral-mediated gene transfer. HSV1-tk gene transduction of one of preestablished wild-type W256 tumor in each animal was accomplished by direct intratumoral injection of retroviral vector-producer cells (W256-->W256TK* tumors). Tumors produced from W256 and W256TK+ cells served as the negative and positive control in each animal. Highly specific images of [124I]-FIAU-derived radioactivity were obtained in W256TK* tumors (that were transduced in vivo) and in W256TK+ tumors but not in nontransduced wild-type W256 tumors. The level of "specific" incorporated radioactivity in transduced portions of both W256TK* and W256TK+ tumors was relatively constant between 4 and 50 h. In the second set, we tested whether [124I]-FIAU and PET imaging can measure and discriminate between different levels of HSV1-tk gene expression. Multiple s.c. tumors were produced from wild-type RG2 cells and stably transduced RG2TK cell lines that express different levels of HSV1-tk. A highly significant relationship between the level of [124I]-FIAU accumulation [% injected dose/g and incorporation constant (Ki)] and an independent measure of HSV1-tk expression (sensitivity of the transduced tumor cells to ganciclovir, IC50) was demonstrated, and the slope of this relationship was defined as a sensitivity index. We have demonstrated for the first time that highly specific noninvasive images of HSV1-tk expression in experimental animal tumors can be obtained using radiolabeled 2'-fluoro-nucleoside [124I]-FIAU and a clinical PET system. The ability to image the location (distribution) of gene expression and the level of expression over time provides new and useful information for monitoring clinical gene therapy protocols in the future.

Midazolam changes cerebral blood flow in discrete brain regions: an H2(15)O positron emission tomography study.

BACKGROUND: Changes in regional cerebral blood flow (rCBF) determined with H2(15)O positron emission tomographic imaging can identify neural circuits affected by centrally acting drugs. METHODS: Fourteen volunteers received one of two midazolam infusions adjusted according to electroencephalographic response. Low or high midazolam effects were identified using post-hoc spectral analysis of the electroencephalographic response obtained during positron emission tomographic imaging based on the absence or presence of 14-Hz spindle activity. The absolute change in global CBF was calculated, and relative changes in rCBF were determined using statistical parametric mapping with localization to standard stereotactic coordinates. RESULTS: The low-effect group received 7.5 +/- 1.7 mg midazolam (serum concentrations, 74 +/- 24 ng/ml), and the high-effect group received 9.7 +/- 1.3 mg midazolam (serum concentrations, 129 +/- 48 ng/ml). Midazolam decreased global CBF by 12% from 39.2 +/- 4.1 to 34.4 +/- 6.1 ml x 100 g(-1) x min(-1) (P < 0.02 at a partial pressure of carbon dioxide of 40 mmHg). The rCBF changes in the low-effect group were a subset of the high-effect group. Decreased rCBF (P < 0.001) occurred in the insula, the cingulate gyrus, multiple areas in the prefrontal cortex, the thalamus, and parietal and temporal association areas. Asymmetric changes occurred, particularly in the low-effect group, and were more significant in the left frontal cortex and thalamus and the right insula. Relative rCBF was increased in the occipital areas. CONCLUSION: Midazolam causes dose-related changes in rCBF in brain regions associated with the normal functioning of arousal, attention, and memory.

Pharmacokinetics and acute toxicology of intraventricular 131 I-monoclonal antibody targeting disialoganglioside in non-human primates.

Tumors metastatic to the leptomeninges are often incurable despite current aggressive treatment modalities. Regional therapy by intrathecal administration of monoclonal antibodies (MoAbs) can maximize their concentration to tumor sites while reducing systemic toxicities. Anti-GD2 antibody 3F8 has successfully targeted human neuroectoderm derived tumors. Disialoganglioside GD2 expression in the central nervous system is identical between humans and cynomolgus monkeys. We studied the pharmacokinetics and the acute and subacute toxicities of intraventricular 131I-3F8 in 8 cynomolgus monkeys. Four animals were purposely immunized with intravenous 3F8 administered 2-4 weeks prior to injections. All animals remained clinically stable. Toxicities included weight loss, fever and CSF leukocytosis. One animal developed a left-sided hemiparesis following his seventh injection, presumably due to a local drug accumulation in the setting of an intermittently patent catheter. The estimated radiation dose to the CSF was 19-48 Gy in the immunized monkeys and 19-82 Gy in the nonimmunized monkeys, and to blood was 0.11-0.98 Gy and 0.29-2.03 Gy, respectively. Histopathology revealed chronic reactive changes adjacent to the region of catheter placement and a focal vasculitis in 2 animals. Peripheral blood counts and bone marrow examinations remained normal. Because of the blood-brain barrier, CSF monkey-anti-mouse antibody titers were less than 10 per cent of those in the serum. In contrast to the CSF radioactivity clearance which was similar in all animals, blood clearance was substantially accelerated in 3F8-immunized animals versus controls. Correspondingly, the CSF to blood dose ratio was improved 1.3 to 6.6 fold (mean 3.5). We conclude that intraventricular administration of 131I-3F8 in primates is tolerable. It can deliver very high doses of radiation to the CSF space with minimal toxicity to blood and bone marrow. Serum anti-mouse antibody accelerates the clearance of 131I-3F8 in blood and may improve the therapeutic index.

Application of laser Doppler flowmetry in neurosurgery.

Experience with LDF has indicated that intraoperative estimation of cerebral blood flow can be very useful and that long-term recordings of cortical perfusion pressure are possible. The disadvantage of LDF is that measurements are extremely local and that the units of measurements are arbitrary. Moreover, the reliability of the technique depends to a large extent on probe position and on the avoidance of movement artifact. The disadvantages are, however, compensated by an excellent dynamic resolution with ultra-short time responses to sudden fluctuations in cerebral perfusion pressure. The high temporal resolution of LDF provides the opportunity to detect hemodynamic events and to monitor the microcirculatory effects of treatment that alters cerebral perfusion pressure in patients with raised intracranial pressure. At no point in the near future is LDF going to supercede continuous recording of intracranial pressure and mean arterial blood pressure and, hence, cerebral perfusion pressure. Rather, we envision LDF becoming an integral part of a multimodal patient monitoring system whereby time trends of monitored variables are combined with calculated variables to provide continuous assessment of cerebral hemodynamic and compensatory reserves.

Infrared laser Doppler flowmeter in the determination of small bowel perfusion after ischemic injury: comparison with the clearance of locally generated hydrogen and fluorescein angiography.

The determination of small bowel perfusion after an ischemic insult is difficult. Regional perfusion was determined in an animal model of neonatal intestinal ischemia using the techniques of laser Doppler flowmetry and the clearance of locally generated hydrogen. Both methods reliably measured tissue perfusion in the areas of maximal ischemic injury. However, considerable variability, perhaps owing to motion artifact, was seen in areas of patchy necrosis. The results suggest that the laser Doppler flowmeter is a suitable technique to measure tissue perfusion in areas of maximal ischemia. However, efforts to reduce motion artifact will be necessary if the laser Doppler is to be used in tissue sites where blood flow is critical for safe anastomosis. This will be a subject of future study.

Comparison of transcutaneous oximetry and laser Doppler flowmetry as noninvasive predictors of wound healing after excision of extremity soft-tissue sarcomas.

BACKGROUND: We wished to determine whether transcutaneous oximetry or laser Doppler flowmetry (LDF) could identify patients at risk for wound failure after conservative, limb-sparing surgery for extremity sarcomas. METHODS: Studies were performed on postoperative days (PODs) 1, 4/5, 7, and 9. Measurements of transcutaneous oxygen pressure (tcPO2) were taken at breathing room air (BL) and 100% oxygen (rate tcPO2). LDF measurements were taken at multiple sites along the wound, and a perfusion index was calculated. RESULTS: Twenty-four patients were studied. Four (17%) had nonhealing wounds. There was no difference in tcPO2 (BL) values between healed and nonhealing wounds. Measurement of rate tcPO2 on POD 1 was significantly lower in the nonhealing wounds than in those with normal healing (28.5 +/- 12.1 mm Hg vs 14.3 +/- 16.2 mm Hg, mean +/- SD, p = 0.03). Rate tcPO2 values increased significantly in healing wounds from POD 1 to PODs 7 and 9 (p = 0.006, p = 0.009). This increase was absent in nonhealing wounds. A clear separation was noted in rate tcPO2 values between groups, with a minimum rate tcPO2 value recorded in a healed wound of 9 mm Hg/min, compared with the maximum value in a nonhealing wound of 7 mm Hg/min. The LDF perfusion index failed to predict wound healing at any of the measured time points. CONCLUSIONS: This study showed that measurement of tcPO2 during oxygen inhalation can accurately predict wound healing in patients after excision of an extremity sarcoma.

Characterization of neuroblastoma xenograft in rat flank. I. Growth, interstitial fluid pressure, and interstitial fluid velocity distribution profiles.

The growth, interstitial fluid pressure (IFP) and interstitial fluid velocity (IFV) profiles of a human neuroblastoma propagated in the flank of an immune suppressed rat were characterized. IFP was measured in the tumor center as a function of size, while radial distributions of IFP and IFV were measured in 2-cm tumors. IFP and IFV were measured using the wick-in-needle and clearance of locally generated hydrogen techniques, respectively. These techniques have a high spatial resolution, permit repetitive measurements, and are minimally invasive. We observed that IFP in the neuroblastoma increased as the tumor grew. Furthermore, IFP increased and its IFV decreased from the periphery toward the center of the tumor. Measured IFP and IFV values were compared to theoretical expectations calculated from the Baxter and Jain mathematical model. The predictions were highly correlated to the measured IFP and IFV profiles with the transport impedence parameter alpha 2 = 24.4. From our measurement data and the Baxter-Jain equations, we computed the interstitium hydraulic conductivity for neuroblastoma to be 7.11 x 10(-6) cm2/mm Hg-sec.

Development of a method to measure kinetics of radiolabelled monoclonal antibody in human tumour with applications to microdosimetry: positron emission tomography studies of iodine-124 labelled 3F8 monoclonal antibody in glioma.

We present a method to assess quantitatively the immunological characteristics of tumours using radiolabelled monoclonal antibody and positron emission tomography (PET) to improve dosimetry for radioimmunotherapy. This method is illustrated with a glioma patient who was injected with 96.2 MBq of iodine-124 labelled 3F8, a murine antibody (IgG3) specific against the ganglioside GD2. Serial PET scans and plasma samples were taken over 11 days. A three-compartment model was used to estimate the plasma to tumour transfer constant (K1), the tumour to plasma transfer constant k2, the association and dissociation constants (k3, k4) of antibody binding, and the binding potential. Tumour radioactivity peaked at 18 h at 0.0045% ID/g. The kinetic parameters were estimated to be: K1 = 0.048 ml h-1 g-1, k2 = 0.16 h-1, k3 = 0.03 h-1, k4 = 0.015 h-1 and BP = 2.25. Based on these kinetic parameters, the amount of tumour-bound radiolabelled monoclonal antibody was calculated. This method permits estimates of both macrodosimetry and microdosimetry at the cellular level based on in vivo non-invasive measurement.

Measurement of brain tissue specific gravity using pycnometry.

In this paper we introduce and characterize pycnometry, a method used to measure fluid density, for determining a tissue's specific gravity. It uses a 2-ml glass pycnometer filled with distilled water to determine a tissue sample's displacement volume. The tissue's density is determined when it's weight is divided by this volume and specific gravity is computed by dividing the tissue density by the density of water. Pycnometry was validated using pre-calibrated glass, specific gravity standards over the range 1.03-1.26, and compared to the density gradient method using rat brain tissue. We observed that the specific gravity values obtained using pycnometry were highly correlated with the specific gravity standards (slope = 1.0107, r = 0.996) and with the density gradient column when tissue volumes larger than 0.120 ml were used with the pycnometer (slope = 1.0707, r = 0.9826). Good correlation was also observed between percent water content values computed using the Nelson equation with pycnometry or density gradient specific gravity values versus the measured percent water content values obtained with the wet weight/dry weight method. Pycnometry is an accurate, reproducible technique to measure tissue specific gravity and brain edema and is best suited for use in a laboratory that engages sporadically in brain edema measurement.

Tumor imaging with carbon-11 labeled alpha-aminoisobutyric acid (AIB) in patients with malignant melanoma.

The potential usefulness of [C-11]-labeled alpha-aminoisobutyric acid (AIB) for tumor imaging has been demonstrated previously in our findings of increased tumor uptake with C-14-labeled AIB in human melanoma heterotransplants in nude mice, and subsequently, in a single case study using C-11 AIB to demonstrate the extent of metastases in patient with widespread malignant melanoma. We report here on the use of C-11 AIB in ten patients with metastatic or unresectable malignant melanoma. Five patients had intense tracer uptake at all known sites of tumor involvement. A sixth patient had good uptake in metastatic lesions in the shoulder and the pelvis, but did not demonstrate uptake within metastatic lesions in the lungs. Two patients had only minimal uptake over the tumor lesions while the two other patients had essentially normal studies. Further studies with C-11 AIB in patients with melanoma and other tumors are warranted.

Cerebrovasculr response to CO2 in edematous brain during either fentanyl or isoflurane anesthesia.

Cerebral blood flow response to changes in PaCO2 was studied in the edematous cerebral cortex of 19 patients with malignant supratentorial tumors using laser Doppler flowmetry technology. General anesthesia for craniotomy was induced with thiopental, 3-5 mg/kg i.v., and N2O, 60% in O2. In random sequence, 8 patients were assigned to receive fentanyl, 6 +/- 1.6 (SEM). mug/kg i.v.; the other 11 received isoflurane, 0.56% end-tidal + 0.07 (SEM). After a craniotomy bone flap was turned and the dura was opened, laser flowmetry probes were placed over surgically undisturbed cortex that was known to be edematous from preoperative CT and MRI scans. Flow index measurements were first made at hypocarbia (PaCO2 = 24.2 +/- 0.9 and 21.5 +/- 2.1 mm Hg for the fentanyl and isoflurane groups, respectively). Minute ventilation was then decreased and cortical flow index was remeasured with PaCO2 = 34.2 +/- 0.6 and 33.0 +/- 0.8 mm Hg for the fentanyl and isoflurane groups, respectively. Hypocarbia during fentanyl-supplemented N2O-O2 anesthesia resulted in a cortical flow index that was 70 +/- 8% of the flow index at near normocarbia (p <0.05). During isoflurane N2O-O2 anesthesia, however, there was a wide variety of responses to hypocarbia, including three patients whose flow indices increased markedly. The mean flow index during hypocarbia was significantly (p <0.05) lower during fentanyl-N2O anesthesia than it was during isoflurane-N2O anesthesia. There was no predictable relationship between the type of brain tumor and the CBF response to hypocapnia during isoflurane-N2O anesthesia. It is concluded that, in edematous brain, cerebral cortical blood flow response to hypocarbia is more likely to be preserved during fentanyl-supplemented N2O-O2 anesthesia than it is during isoflurane-supplemented N2O-O2 anesthesia. In neuropathologic states where hyperventilation is thought to be necessary to reduce cerebral blood flow and decrease brain bulk, isoflurane may be less satisfactory than fentanyl as a supplement to N2O-O2 anesthesia.

The effect of the aminosteroid U-78517G on peritumoural brain oedema.

U-78517G belongs to a novel group of compounds-aminosteroids (AS) which are potent inhibitors of central nervous system tissue lipid peroxidation and are devoid of classical glucocorticoid and mineralocorticoid activities. The AS have been found to possess cerebral protective properties against ischaemia, ischaemic oedema, trauma to the cerebrum and spinal cord and in sub-arachnoid hemorrhage. The possible efficacy of U-78517G in attenuating peritumoural oedema was examined in rats harbouring cerebral 9L gliosarcomas. 19 Sprague-Dawley rats were implanted intracranially with 9L gliosarcoma cells. On day 14 post implantation rats were randomized to control (no treatment n = 10) and AS treated animals (n = 9). On day 20 animals were sacrificed and six brain samples per rat were examined for the percentage of water content. Oedema was assessed with the wet/dry weight technique. The AS treated animals were found to have a significant increase in peritumoural oedema (+1.38%, P less than 0.001) possibly related to an increase in cerebral blood flow associated with AS treatment. Further studies are, however, underway to precisely clarify this unexpected observation.

Measurement of brain tissue density using pycnometry.

A novel method to measure specific gravity (SG) of tissues, pycnometry (PYC), is described. This method utilizes a 2ml glass pycnometer filled with distilled H2O to determine the displacement volume of a tissue sample and an equation to compute SG from the sample's weight and the pycnometer's weight before and after adding the sample. The PYC method was validated using glass SG standards over the range 1.02-1.26, and against the column density gradient (DG) method using brain tissue from 250-300 g male rats. Factors which affect PYC accuracy, i.e. sample size, were also evaluated. Our results indicate that PYC SG values are highly correlated with the glass SG standards (slope = 1.0107, r = 0.9955, p less than 0.001), and highly correlated with DG when approximately 0.120 ml tissue samples are used in the pycnometer. The DG method was preferable to the PYC method, however, when small tissue samples, i.e. 0.60 ml or less, were used.

Intraoperative measurement of cerebral and tumor blood flow with laser-Doppler flowmetry.

A new technique, laser-Doppler flowmetry, has been used intraoperatively to measure blood flow responses in normal brain tissue and brain tumor to blood pressure and arterial blood gas alterations. We have observed that blood flow is reduced in most cerebral tumors, and that most tumors retain the normal response to changes in arterial blood gas; however, these responses are varied. One group of tumors in our study demonstrated an autoregulatory capacity; a second behaved passively--that is, blood flow changes followed blood pressure--while a third showed no response.

Validation of transport measurements in skeletal muscle with N-13 amino acids using a rabbit isolated hindlimb model.

We are studying the transport of C-11 and N-13 labeled amino acids in tumor-bearing rabbits to determine the role of amino acid transport in the pathogenesis of muscle wasting in cancer. To validate a new, in vivo, method for measuring transport in skeletal muscle with these compounds, an isolated hindlimb model was developed in rabbits. The limb was perfused with a non-recirculating, normothermic, constant pressure system and a cell-free perfusate. Hemodynamic and metabolic parameters were measured during the first 75 min. of perfusion and found to remain normal and stable. Flow varied directly with perfusion pressure over the normal range of resting flows in the intact rabbit hindlimb. Time-activity curves (TAC's) were recorded from the medial thigh following bolus co-injection of L-[amide N-13] glutamine or N-13 L-glutamate with Tc-99m human serum albumin (HSA) into the femoral artery. Regional plasma flow was determined from the Tc-99m data. The N-13 TAC's consistently manifested a three-phased washout with half times of approximately 30 sec., 5 min. and 2 hr. Capillary and cellular transport parameters were computed from the N-13 data using a double barrier, single capillary model of capillary and cellular transport and assuming that the three washout components result, respectively, from tracer throughput, extraction into the interstitial space and extraction into the intracellular space. This interpretation was validated and the sensitivity of the technique to transport processes demonstrated by examining the effects on the N-13 TAC's and computed transport parameters of several factors known to influence cellular transport of amino acids, viz., the insulin concentration, amino acid concentration and pH of the perfusate. Time-activity curves and transport parameters for N-13 L-glutamine in the isolated limb were very similar to those observed in the intact rabbit hindlimb, suggesting that studies in the perfused model are indicative of amino acid transport in vivo. The methodology described here is especially well suited for studying the specific effects on transport of factors which influence amino acid metabolism in skeletal muscle (e.g., hormones and monokines).