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K Wasserman

Publications and source records attributed to K Wasserman.

At least 73 records · Page 4Linked to original sources
PubMed1994

A method for estimating bicarbonate buffering of lactic acid during constant work rate exercise.

A method to estimate the CO2 derived from buffering lactic acid by HCO3- during constant work rate exercise is described. It utilizes the simultaneous continuous measurement of O2 uptake (VO2) and CO2 output (VCO2), and the muscle respiratory quotient (RQm). The CO2 generated from aerobic metabolism of the contracting skeletal muscles was estimated from the product of the exercise-induced increase in VO2 and RQm calculated from gas exchange. By starting exercise from unloaded cycling, the increase in CO2 stores, not accompanied by a simultaneous decrease in O2 stores, was minimized. The total CO2 and aerobic CO2 outputs and, by difference, the millimoles (mmol) of lactate buffered by HCO3- (corrected for hyperventilation) were estimated. To test this method, ten normal subjects performed cycling exercise at each of two work rates for 6 min, one below the lactic acidosis threshold (LAT) (50 W for all subjects), and the other above the LAT, midway between LAT and peak VO2 [mean (SD), 144 (48) W]. Hyperventilation had a small effect on the calculation of mmol lactate buffered by HCO3- [6.5 (2.3)% at 6 min in four subjects who hyperventilated]. The mmol of buffer CO2 at 6 min of exercise was highly correlated (r = 0.925, P < 0.001) with the increase in venous blood lactate sampled 2 min into recovery (coefficient of variation = +/- 0.9 mmol.l-1). The reproducibility between tests done on different days was good. We conclude that the rate of release of CO2 from HCO3- can be estimated from the continuous analysis of simultaneously measured VCO2, VO2, and an estimate of muscle substrate.

Adult↗
PubMed1994

Critical capillary oxygen partial pressure and lactate threshold in patients with cardiovascular disease.

OBJECTIVES: The aim of this study was to determine the relation between femoral vein oxygen partial pressure (PO2) and lactate increase during exercise in patients with cardiovascular disease. BACKGROUND: Considerable controversy surrounds the relation between the increase in lactate during exercise and the oxygen supply to the exercising muscles. We assumed that femoral vein PO2 would be a measure of end-capillary PO2 during leg-cycling exercise and that it would decrease to a "floor" level when the critical capillary PO2 (the PO2 below which the capillary-mitochondrial difference would be too low to allow oxygen consumption) was reached. At the critical capillary PO2, anaerobic metabolism should take place, and lactate should increase in the effluent blood. METHODS: Ten patients with cardiovascular disease performed two 6-min constant work rate tests (moderate and heavy intensity) and an incremental exercise test to the symptom-limited maximum on a cycle ergometer. Femoral vein blood was repeatedly sampled through a percutaneous catheter before and during each exercise test. RESULTS: The PO2 rapidly decreased toward a minimal value with increasing oxygen uptake for all three tests in all patients. After reaching its nadir (18.2 +/- 2.0 mm Hg), the PO2 remained unchanged in five patients but increased in the other five patients despite the further increase in work rate and oxygen uptake. The relation between PO2 and oxygen uptake was characteristic for each patient and independent of the protocol used for the study. Femoral vein lactate did not change appreciably until PO2 reached the minimal (critical) value. Thereafter, it dramatically increased without a further decrease in PO2. The minimal PO2 was positively correlated with the peak oxygen uptake (r = 0.70, p = 0.01). CONCLUSIONS: During leg-cycling exercise, muscle capillary PO2 reaches a minimal value in the midrange of the subjects' work capacity before lactate concentration increases in patients with cardiovascular disease. The lack of further decrease in PO2 at the oxygen uptake at which lactate starts to increase suggests that the minimal capillary PO2 is the "critical" capillary PO2.

Aged↗
PubMed1994

Coupling of external to cellular respiration during exercise: the wisdom of the body revisited.

The changes in cellular respiration needed to increase energy output during exercise are intimately and predictably linked to external respiration through the circulation. This review addresses the mechanisms by which lactate accumulation might influence O2 uptake (VO2) and CO2 output (VCO2) kinetics. Respiratory homeostasis (a steady state with respect to VO2 and VCO2) is achieved by 3-4 min for work rates not associated with an increase in arterial lactate. When blood lactate increases significantly above rest for constant work rate exercise, VO2 characteristically increases past 3 min (slow component) at a rate proportional to the lactate concentration increase. The development of a similar slow component in VCO2 is not evident. The divergence of VCO2 from VO2 increase can be accounted for by extra CO2 release from the cell as HCO3- buffers lactic acid. Thus the slow component of aerobic CO2 production (parallel to VO2) is masked by the increase in buffer VCO2. This CO2, and the consumption of extracellular HCO3- by the lactate-producing cells, shifts the oxyhemoglobin dissociation curve rightward (Bohr effect). The exercise lactic acidosis has been observed to occur after the minimal capillary PO2 is reached. Thus the lactic acidosis serves to facilitate oxyhemoglobin dissociation and O2 transport to the muscle cells without a further decrease in end-capillary PO2. From these observations, it is hypothesized that simultaneously measured dynamic changes in VO2 and VCO2 might be useful to infer the aerobic and anaerobic contributions to exercise bioenergetics for a specific work task.

Acidosis, Lactic↗
PubMed1994

Lactic acidosis as a facilitator of oxyhemoglobin dissociation during exercise.

The slow rise in O2 uptake (VO2), which has been shown to be linearly correlated with the increase in lactate concentration during heavy constant work rate exercise, led us to investigate the role of H+ from lactic acid in facilitating oxyhemoglobin (O2Hb) dissociation. We measured femoral venous PO2, O2Hb saturation, pH, PCO2, lactate, and standard HCO3- during increasing work rate and two constant work rate cycle ergometer exercise tests [below and above the lactic acidosis threshold (LAT)] in two groups of five healthy subjects. Mean end-exercise femoral vein blood and VO2 values for the below- and above-LAT square waves and the increasing work rate protocol were, respectively, PO2 of 19.8 +/- 2.1 (SD), 18.8 +/- 4.7, and 19.8 +/- 3.3 Torr; O2 saturation of 22.5 +/- 4.1, 13.8 +/- 4.2, and 18.5 +/- 6.3%; pH of 7.26 +/- 0.01, 7.02 +/- 0.11, and 7.09 +/- 0.07; lactate of 1.9 +/- 0.9, 11.0 +/- 3.8, and 8.3 +/- 2.9 mmol/l; and VO2 of 1.77 +/- 0.24, 3.36 +/- 0.4, and 3.91 +/- 0.68 l/min. End-exercise femoral vein PO2 did not differ statistically for the three protocols, whereas O2Hb saturation continued to decrease for work rates above LAT. We conclude that decreasing capillary PO2 accounted for most of the O2Hb dissociation during below-LAT exercise and that acidification of muscle capillary blood due to lactic acidosis accounted for virtually all of the O2Hb dissociation above LAT.

Acidosis, Lactic↗
PubMed1994

Evaluation of a symmetrically disposed Pitot tube flowmeter for measuring gas flow during exercise.

We evaluated the effect of airflow and gas composition on the linearity of measurement of airflow by a new disposable flowmeter. The flowmeter is based on the principle of differential pressure measurement across two symmetrically disposed Pitot tubes. Nonlinearities arising from the pressure-to-airflow relationship and sensitivity to changes in gas density were linearized with appropriate software and monitoring of the gas composition. With room air used as the respired gas, the measured tidal volume from a piston pump assembly was consistently within 1-2% of the target tidal volume for each of five flowmeters tested across physiological ranges of flow. Changing gas densities by varying concentrations of O2, CO2, and N2 led to errors in tidal volume measurement that ranged up to 6-8%. However, because the errors were predictable, they were corrected by software to within 0.6% of the target volume. Measurement of minute ventilation during exercise was within 1-2% of that determined from bag collections. We conclude that this type of flowmeter can accurately measure exercise minute ventilation and has advantages over some other flowmeters because of its ruggedness, reproducibility, and ease of sterilization or replacement compared with other flowmeters.

Equipment Design↗
PubMed1994

Expression of surface markers on alveolar macrophages from symptomatic patients with HIV infection as detected by flow cytometry.

Alveolar macrophages (AMs) harvested from 32 HIV-infected patients with respiratory problems (opportunistic pulmonary infections, n = 12; other lung disease, n = 20) and 13 healthy controls were stained with a panel of 15 monoclonal antibodies directed against surface antigens implicated in cell function. Antigen expression was quantified by flow cytometry and expressed as relative linear median fluorescence intensity (RLMFI). On AMs of patients, as compared with controls, there was a significant enhancement of HLA DP (12.1 +/- 1.5 vs 6.5 +/- 0.9, p = 0.01, M +/- SEM, RLMFI units), CD11b (3.4 +/- 0.5 vs 1.7 +/- 0.4, p = 0.014), CD11c (8.9 +/- 1.0 vs 4.8 +/- 0.8, p = 0.0046), CD14 (2.1 +/- 0.3 vs 1.0 +/- 0.2, p = 0.0009), and CD33 (1.7 +/- 0.1 vs 1.0 +/- 0.2, p = 0.0093). No significant differences could be established for HLA-DR (36.9 +/- 5.8 vs 30.9 +/- 7.5, NS), HLA-DQ (3.4 +/- 0.3 vs 3.1 +/- 0.6, NS), CD54 (1.9 +/- 0.3 vs 1.2 +/- 0.1, NS), CD13 (2.5 +/- 0.6 vs 1.5 +/- 0.3, NS), CD36 (1.4 +/- 0.2 vs 0.9 +/- 0.3, NS), CD71 (10.3 +/- 1.9 vs 8.9 +/- 1.8, NS), CD25 (0.8 +/- 0.0 vs 0.9 +/- 0.1, NS), 27E10 (1.1 +/- 0.1 vs 0.8 +/- 0.3, NS), RM3/1 (1.9 +/- 0.4 vs 1.5 +/- 0.4, NS), and CD4 (1.5 +/- 0.3 vs 1.0 +/- 0.0, NS). The expression of CD14 and CD11b, but not of HLA class II antigens and CD71, was increased in the smaller cell population compared with the larger, thus suggesting monocyte recruitment. The increased expression of HLA-DP, CD11c, CD14, and CD33 on the patients' AMs was independent of smoking habits. The degree of immunodeficiency as indicated by the absolute peripheral CD4 count, the character of HIV-related pulmonary disease, and the prophylactic use of pentamidine or zidovudine did not significantly modify the antigen expression pattern. It is concluded that HIV infection may lead, most probably indirectly, to enhanced expression of surface antigens by local upregulation and/or recruitment of monocytes from the peripheral circulation. The functional significance of enhanced marker expression requires further clarification.

Adult↗
PubMed1994

Dynamics of oxygen uptake for submaximal exercise and recovery in patients with chronic heart failure.

STUDY DESIGN AND OBJECTIVES: Attainment of a steady state for oxygen uptake (VO2) during constant work rate exercise has been reported to take longer for patients with chronic heart failure (CHF) compared with normal. The steady state is also delayed in normal subjects during high-intensity exercise compared with moderate exercise, however, and the delay correlates with the degree of associated lactic acidosis. To determine whether prolonged kinetics of VO2 are attributable solely to the reduction of exercise capacity in CHF, VO2 kinetics were compared for patients with CHF and normal subjects, both for exercise of matched absolute work rate and for matched relative work intensity. SUBJECTS: Eighteen men with CHF and 10 normal men. METHODS AND RESULTS: Subjects performed 6 min of constant work rate cycle ergometry with breath-by-breath measurement of VO2. Patients were studied using 25 W, and a work rate midway between the lactic acidosis threshold and maximal capacity (50 percent delta). Normal subjects were tested similarly, and also at a work rate matched to the patients' average 50 percent delta work rate. The VO2 kinetics were characterized by the mean response time (MRT) to attain the 6 min VO2 value. Rates of recovery of VO2 were analyzed for 2 min following exercise. For the same absolute work rate, VO2 MRTs were significantly longer for patients than controls (25 W, 67 +/- 26 vs 37 +/- 25 s; approximately 60 W, 87 +/- 20 vs 54 +/- 27 s), but there was no significant difference in VO2 MRT between the two groups at a matched intensity of 50 percent delta (87 +/- 20 vs 81 +/- 18 s). However, the decrease in VO2 during 2 min of recovery was slower for the patients on all comparisons, even for matched exercise intensity. CONCLUSION: The VO2 dynamics for submaximal exercise are slowed in CHF. The slower dynamics are not entirely accounted for by the relatively higher intensity of a given work rate, since delayed recovery is evident even at a matched relative work intensity. Exercise intolerance in CHF is characterized not only by decreased maximal exercise capacity, but also by slower adaptations to and from submaximal levels of exercise.

Anaerobic Threshold↗
PubMed1994

Determination of the anaerobic threshold by gas exchange: biochemical considerations, methodology and physiological effects.

This paper explains the physiological and biochemical basis of the anaerobic threshold (AT), achieved during physical exercise. The lactate concentration is approximately the same at rest in relatively fit adults, in normal sedentary subjects in adult patients with heart disease. But during exercise, the increase of lactate is inversely related to the physical fitness of the individual. During incremental work, the lactate concentration increases initially very little until a distinct metabolic rate (VO2 AT) is reached at which lactate starts to increase steeply (anaerobic threshold/AT; VO2 AT). Above the anaerobic threshold, accelerated glycolysis increases muscle lactic acidosis. This acidosis is buffered primarily by bicarbonate. The bicarbonate-derived CO2 causes an increased alveolar CO2 output relative to O2 uptake. Oxygen uptake is increased virtually linearly with work rate in healthy subjects with a slope of approximately 10 ml O2/min/Watt. VCO2 starts to increase more steeply in the mid-work-rate range after an initial linear behavior. This steepening is caused by an increased CO2 production from the HCO3-buffering of lactic acid for the range of work rates above the AT. Below the AT, the slope of increase in VCO2 is 1 or slightly less, averaging 0.95. Above the AT, it is greater than 1. The submaximal exercise protocol for the determination of AT includes a period of 2-3 min of unloaded cycling, a ramp program with x Watt increase/minute and a recovery period of 2 min. X is the rate of work rate increase per min, so that the incremental period of the exercise test lasts 8-10 min, stressing the patient for only a short time. The anaerobic threshold can be determined during the ramp program using the following four parameters: 1) steeper increase of VCO2 as compared to VO2 (V-slope-method); 2) respiratory exchange ratio = 0.95; 3) PETO2 increase; 4) VE/VO2 increase. The V-slope-method can be successfully applied, not only in healthy volunteers, but also in patients suffering from cardiac and/or pulmonary (breathing abnormalities) diseases. The so far published data show that the anaerobic threshold in healthy people and patients is a highly reproducible, accurately measurable, securely achievable parameter for the non-invasive evaluation of the individual cardiopulmonary exercise capacity.

Acid-Base Equilibrium↗
PubMed1993

Femoral vascular occlusion and ventilation during recovery from heavy exercise.

Ventilation and cardiac output subside gradually following cessation of exercise, which is commonly linked to the slow wash-out of materials from the recovering muscles. The effect of hindering the removal of the metabolic products of heavy cycle exercise on the kinetics of ventilation and gas exchange was studied in 5 subjects by occluding the femoral circulation with cuffs during the first 2 min of recovery (15 tests). Fifteen undisturbed recoveries served as controls. Compared to spontaneous recovery, circulatory obstruction induced an immediate (from the first breath) decrease in minute ventilation (VE), while end-tidal CO2 (PETCO2) as well as lactate and K+ in venous blood at forearm did not change significantly. A ventilatory deficit of 27 +/- 9 L was observed from the 2 min of occlusion. Following cuff deflation, VE rose 2-3 breaths after PETCO2 began to increase in every subject. The mechanisms of the normocapnic reduction of VE during occlusion, as well as the rise of ventilation following cuff release, are still unclear. However, these results argue against any significant role for hyperpnea-inducing intramuscular chemoreception, or point to muscular perfusion as a prerequisite of such a mechanism to operate.

Adult↗
PubMed1993

Role of muscle perfusion and baroreception in the hyperpnea following muscle contraction in dog.

The influence of impeding muscle perfusion on the time course of ventilatory decline during recovery from electrically induced hindlimb contractions has been studied in 14 anesthetized dogs. When intravascular balloons, placed in abdominal aorta and inferior vena cava just rostral to the iliac bifurcation, were inflated at the cessation of contraction bout, minute ventilation (VE) was significantly reduced during recovery compared with control. The subsequent restoration of iliac circulation rapidly augmented VE, which peaked at the fifth breath after release, by an average of +4.97 L.min-1; VE then returned exponentially to resting (pre-contraction) level. Breathing 100% O2 did not affect the VE recovery pattern neither during iliac occlusion nor immediately after its release (the peak average delta VE = +4.42 L.min-1). When a local anesthetic (5% Lidocaine) was applied bilaterally to the regions of carotid bifurcation, systemic blood pressure was significantly increased and the VE response to both iliac occlusion and release were nearly abolished. The VE response to inhalation of 5% CO2 in air was not affected by this procedure, whereas the stimulation of VE with 2 mg i.v. bolus of NaCN was attenuated. When the local anesthetic was thoroughly washed out (and systemic blood pressure had returned to control level) the previously observed VE responses to iliac occlusion and release were restored. These results and analysis of the VE response timing (transits and latencies) suggest that the vascular rather than humoral effects or tissue 'metaboreception' modulate ventilatory recovery from muscular contractions; baroreception appears to be important in this process.

Animals↗
PubMed1993

O2 uptake kinetics and the O2 deficit as related to exercise intensity and blood lactate.

The dynamic responses of O2 uptake (VO2) to a range of constant power output levels were related to exercise intensity [as percent maximal VO2 and as below vs. above lactic acid threshold (LAT)] and to the associated end-exercise lactate in three groups of subjects: group I, untrained subjects performing leg cycle ergometer exercise; group II, the same subjects performing arm cycle exercise; and group III, trained cyclists performing leg cycle ergometer exercise. Responses were described by a double-exponential equation, with each component having an independent time delay, which reduced to a monoexponential description for moderate (below-LAT) exercise. When a second exponential component to the VO2 response was present, it did not become evident until approximately 80-100 s into exercise. An overall time constant (tau T, determined as O2 deficit for the total response divided by net end-exercise VO2) and a primary time constant (tau P, determined from the O2 deficit and the amplitude for the early primary VO2 response) were compared. The tau T rose with power output and end-exercise lactate levels, but tau P was virtually invariant, even at high end-exercise lactate levels. Moreover the gain of the primary exponential component (as delta VO2/delta W) was constant across power outputs and blood lactate levels, suggesting that the primary VO2 response reflects a linear system, even at higher power outputs. These results suggest that elevated end-exercise lactate is not associated with any discernible slowing of the primary rise in VO2.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1993

O2 uptake kinetics above and below the lactic acidosis threshold during sinusoidal exercise.

O2 uptake (VO2) kinetics at the onset of a constant work rate exercise are difficult to describe for work rates above the lactic acidosis threshold (LAT), because the steady-state level of VO2 response can usually not be identified. To describe the ability of the O2 transport system to deliver and the cells to utilize O2 above the LAT relative to that below the LAT, we applied a fluctuating (sinusoidal) variation of work rate. After 4 min of constant work at the midpoint of the sinusoidal work rate, a fluctuating work rate, at a period of 4 min, was applied below the LAT for the next 16 min. This was repeated in a range of work rates above the LAT with the same sine-wave amplitude. VO2 response appeared to follow a sinusoidal pattern similar to that of work rate for below- and above-LAT exercise. However, the amplitude of the VO2 response was significantly reduced (5.4 +/- 2.6 vs. 7.6 +/- 1.9 ml.min-1 x W-1, P < 0.01), and the phase lag increased above- compared with below-LAT work rate. VO2/heart rate fluctuations were dramatically reduced, whereas heart rate amplitude decreased and phase lag increased, for above-LAT sinusoidal work rate changes. These results suggest that VO2 kinetics are slowed in the work rate domain above the LAT relative to that below the LAT and that VO2 kinetics could be limited by the O2 transport mechanisms to the exercising muscle.

Acidosis, Lactic↗
PubMed1993

Comparison of gas exchange, lactate, and lactic acidosis thresholds in patients with chronic obstructive pulmonary disease.

During an incremental exercise test, three consequences of the onset of anaerobic metabolism can be observed: rise in blood lactate (lactate threshold, LT); fall in standard bicarbonate (lactic acidosis threshold, LAT); nonlinear increase in CO2 output (V-slope gas exchange threshold, GET). We compared these thresholds in 31 patients with COPD. We found that the GET and LAT overestimated the LT. A better relationship was found between LAT and GET, even though GET was significantly higher than LAT (by 124 ml/min; p < 0.0001). However, since the bias is appreciably greater at lower LAT values (likely because VCO2 kinetics are slower than VO2 kinetics), we separated the studies into two groups: (A) tests where LAT occurred within the first 2 min of the increasing work rate period, and (B) tests where LAT occurred after 2 min. For Group A, there was a substantial bias between LAT and GET (323 ml/min, p < 0.0001), whereas the bias was much smaller (only 5.4%, though statistically significant) for Group B (57 ml/min, p < 0.01). We conclude that when lactic acidosis occurs after the first 2 min of incremental exercise, the GET closely approximates the point at which blood bicarbonate begins to fall.

Acidosis, Lactic↗
PubMed1993

O2 uptake kinetics in response to exercise. A measure of tissue anaerobiosis in heart failure.

Oxygen uptake (VO2) reflects the rate of aerobic regeneration of high-energy phosphate compounds (primarily adenosine triphosphate [ATP]). Since lactate increase is thought to result from an inadequate rate of aerobic ATP regeneration, it might be expected that lactate increase would be associated with a delayed attainment of steady state for VO2 in response to constant load exercise. Similarly if mitochondrial ATP regeneration during exercise is inadequately supported by O2 transport mechanisms, adenosine diphosphate (ADP) and purine nucleotide by-products, such as hypoxanthine, should increase. This study investigated the relationship between VO2 kinetics during exercise and accompanying changes in blood lactate and hypoxanthine values in heart failure patients, as a model of compromised O2 transport. Twenty-five patients with chronic heart failure performed cycle ergometry for 6 min at 25 W and at a work rate midway (50 percent delta) between their lactic acidosis threshold (LAT) and peak VO2. Ventilation and gas exchange were measured breath by breath, and venous lactate, hypoxanthine, norepinephrine, and epinephrine were determined at rest and 2 min after each test. The slow component of VO2 kinetics was quantified as the rise in VO2 from the third to the sixth minute of exercise (delta VO2 [6-3]). Ten age- and size-matched normal subjects served as control subjects. delta VO2 (6-3) was correlated with the increase in lactate (r = 0.71, p < 0.001), hypoxanthine (r = 0.61, p < 0.001), and norepinephrine (r = 0.41, p < 0.01) but not epinephrine in response to exercise in the heart failure patients. The delta VO2 (6-3) and delta lactate were both greater in the patients than in the control subjects at similar absolute work rates (54 +/- 20 and 60 W, respectively). However, the slope of the relationship between delta La and delta VO2 (6-3) for the patient and normal groups was indistinguishable. The lactate increase was correlated with hypoxanthine increase (r = 0.66, p < 0.001), but not norepinephrine or epinephrine. In summary, VO2 kinetics in response to exercise reflects delayed attainment of the steady state in heart failure patients, which is correlated with increases in lactate and hypoxanthine, markers of increased anaerobic metabolism.

Acidosis, Lactic↗
PubMed1993

Uses of oxygen in the treatment of acute respiratory failure secondary to obstructive lung disease.

Breathing 100% O2 is a way of slowing breathing rate, decompressing hyperinflated, low VA/Q lung units, reducing the FRC and allowing the patient with COPD in acute respiratory failure to breath at a more comfortable lung volume and with a higher tidal volume. This should also improve the efficiency of breathing, increasing the fractional turnover of alveolar gas. Simultaneously O2 protects the brain, heart and other organs from hypoxic injury. Breathing 100% O2 provides the major therapeutic approach for rapidly reversing the pathophysiological cycle of impaired respiratory mechanics which threatens the life of the patient in acute respiratory failure. It is a therapeutic approach which might be employed in the tachypnoeic patient in acute respiratory distress, before initiating airway intubation and mechanical ventilation. This provides the physician with time to allow other mechanisms of bronchodilating the patient to be started, and for the need of a more aggressive approach to airway management to be evaluated.

Acute Disease↗
PubMed1992

Nongranular proteolytic enzymes of rat IL-2-activated natural killer cells. I. Subcellular localization and functional role.

Our investigations indicate that a variety of neutral serine proteases exist in highly purified, IL-2-activated rat NK (A-NK) cells. These enzymatic activities are not restricted to only cytolysin-containing granules and are not defined by only the assay of N-alpha-benzyloxycarbonyl-L-lysine thiobenzylesterase activity. These activities, which we term A-NKP 1, A-NKP 2, A-NKP 3, and A-NKP 4, cleave, respectively, the following fluorogenic peptide substrates: Boc-Phe-Ser-Arg-7-amino-4-methylcoumarin (AMC, trypsin-like); Suc-Ala-Ala-Phe AMC (chymotrypsin-like); Suc-Gly-Pro-Leu-Gly-Pro AMC (collagenase-like), and Z-Phe-Arg AMC (another trypsin-like enzyme). The proteases A-NKP 1, A-NKP 2, and A-NKP 3 are not cell surface-associated and appear to be cytosolic as defined by isopycnic sucrose density gradient centrifugation. In contrast, A-NKP 4 appears to be located in lysosomes. Treatment of rat A-NK cells with protease inhibitors that inhibit A-NKP 2 and A-NKP 3 also substantially inhibit A-NK cell-mediated cytotoxicity against both NK-sensitive and -resistant targets (YAC-1 and P815, respectively). These results indicate that A-NKP2 and A-NKP 3 may play a role in IL-2-activated NK cell-mediated cytotoxicity. A variety of proteolytic enzymes, in addition to granzymes, therefore exist in A-NK cells. Our studies indicate that a prerequisite to a thorough understanding of the role of proteases in killer cell function is the investigation of several classes of enzymes in addition to granzymes contained in lytic granules.

Animals↗