JOURNAL
OF SURGICAL
27, 84-92 (1979)
RESEARCH
Impaired Reactivity in Autologous
of Burn Patient Lymphocytes to Phytohemagglutinin Serum: Failure to Improve Responsiveness by Washing In Vitro MARK
Division
B. CONSTANTIAN,
of Plastic Surgery, Medical College Submitted
for publication
M.D.’
of Virginia,
Richmond.
Virginia
23298
August 11, 1978
Twenty-one peripheral blood lymphocyte samples from 17 burned patients with injuries of varying severity were examined for reactivity to stimulation by phytohemagglutinin in pooled and autologous serum, and compared to samples from simultaneously tested normalcontrols. The bum lymphocytes were less reactive than controls (P < 0.05) in either serum, but were most hyporesponsive in autologous serum. The presence of immunosuppressive activity in bum serum did not correlate with the degree of cellular hyporesponsiveness to PHA. The reactivily could not be improved by washing the cells prior to stimulation. These results indicate that the impaired responsiveness of lymphocytes from bum patients to PHA is not due to loose association of a circulating immunosuppressor with the cell membrane and may, instead, be due in part to the emergence of a suppressor T-cell population.
Consistent with this reasoning, there is evidence that the serum from patients who Multiple host immunological deficits folhave undergone bum injury or other trauma low operative or accidental trauma, includdepresses the response of normal human ing bums; among them is a profound impairlymphocytes to mitogenic stimulation [7,20, ment of T-cell dependent reactions [l, 3, 1.5, 241. Our prior reports [6, 71 have indicated 19, 20, 241. Testing by a number of laborathat the serum of operatively or accidentally tories has established that the peripheral traumatized patients suppresses the reblood lymphocytes of traumatized patients sponse of normal lymphocytes to PHA. This are hyporesponsive to stimulation by phytoimmunosuppressive activity parallels in hemagglutinin (PHA) in vitro 125, 26, 281. degree and duration the severity of the This has been taken as evidence of a non- clinical course and appears to be localized specific depression of T-cell reactivity which in a low molecular weight ( ‘0.1) between bum and control cell ratios in either pooled or autologous serum. Examined individually, however, (Tables 4 and 5) it can be seen that while the cells of 4 of 8 controls had a wash index of 2 or greater in autologous serum (and hence were at least twice as reactive after six washes than after one wash), only 1 of 21 burn patients was as reactive. This difference was significant (P < 0.025) by x2 analysis. The differences in pooled serum (4 of 8 controls versus 7 of 21 patients) were not significant (P > 0.5). DISCUSSION
Recent evidence for post-traumatic lymphocyte hyporeactivity, especially to stimuli which require the participation of T-cells
CONSTANTIAN:
BURN
PATIENT
TABLE
89
LYMPHOCYTES
4
RESPONSE OF CELLS WASHED SIX TIMES TO PHA IN A~TOLOGO~S SERUM (cpm f SE) Patients
Controls Experiment No. 8 8 14 14 14 15 15 16 16 17 17 17 18 18 21 22 22 22 23 23 Mean f. SE
No PHA
295 2
PHA (l/200)
9
305 “_ 60 156 2
55
7,169 -c
633
13,952 rc_2,455 1,905 +
395
401 2 105
38,224 2 1,860
695 k 219
4,198 -+ 1,165
220 k 40 1,708 2 573
48,466 a 1,356 6,310 2 2,866
1,118 2
29,836 r
36
612 2 208
590
18,753 2 6,805
(1, 19, 20, 24-26, 28) has been presumed to explain in part the prevalence of lethal infectious complications following burns and other injuries. While this may be the case, the origin and nature of the (lymphocyte) hyporesponsiveness has not been established. Data from a number of laboratories have indicated that the serum of traumatized patients contain immunosuppressive activity which can inhibit antibodydependent cell-mediated cytolysis [30] and the response of peripheral blood lymphocytes to allogeneic cells [20], to Moniliu antigen [24], and to PHA [5, 71. This evidence has suggested that the observed host lymphocyte defects following trauma may be extrinsic and due to potentially reversible cell membrane changes brought about by a serum component. Our prior results indicate that in both surgically traumatized [7] and burned patients [5] a majority of the immunosuppressive serum activity is located in a low molecular
No PHA 138 ” 1.0 158 401 2 19 155 f 8 1,956 k 44 602 + 300 897 2 108 697 + 39 1,430 f 405 582 + 237 1,322 2 418 3,005 +- 184 629 k 267 2,051 2 568 660 k 346 2,033 2 103 589 r+ 51 907 2 80 1,291 4 96 923 5 103 1,198 -r- 252
PHA (l/200) 5,184 k 2,076 10,637 -+ 1,698 706 2 156 820 f 482 1,486 k 51 338 ? 42 1,037 f 37 158 4 44 4,956 2 570 3,681 + 177 3,745 2 947 5,448 c 864 588 2 174 846 -c 409 5,972 + 357 2,406 2 395 9,653 k 1,815 4,438 2 128 15,230 f 2,230 24,383 f 1,279 4,925 4 1,288
weight (
OF SURGICAL
27, 84-92 (1979)
RESEARCH
Impaired Reactivity in Autologous
of Burn Patient Lymphocytes to Phytohemagglutinin Serum: Failure to Improve Responsiveness by Washing In Vitro MARK
Division
B. CONSTANTIAN,
of Plastic Surgery, Medical College Submitted
for publication
M.D.’
of Virginia,
Richmond.
Virginia
23298
August 11, 1978
Twenty-one peripheral blood lymphocyte samples from 17 burned patients with injuries of varying severity were examined for reactivity to stimulation by phytohemagglutinin in pooled and autologous serum, and compared to samples from simultaneously tested normalcontrols. The bum lymphocytes were less reactive than controls (P < 0.05) in either serum, but were most hyporesponsive in autologous serum. The presence of immunosuppressive activity in bum serum did not correlate with the degree of cellular hyporesponsiveness to PHA. The reactivily could not be improved by washing the cells prior to stimulation. These results indicate that the impaired responsiveness of lymphocytes from bum patients to PHA is not due to loose association of a circulating immunosuppressor with the cell membrane and may, instead, be due in part to the emergence of a suppressor T-cell population.
Consistent with this reasoning, there is evidence that the serum from patients who Multiple host immunological deficits folhave undergone bum injury or other trauma low operative or accidental trauma, includdepresses the response of normal human ing bums; among them is a profound impairlymphocytes to mitogenic stimulation [7,20, ment of T-cell dependent reactions [l, 3, 1.5, 241. Our prior reports [6, 71 have indicated 19, 20, 241. Testing by a number of laborathat the serum of operatively or accidentally tories has established that the peripheral traumatized patients suppresses the reblood lymphocytes of traumatized patients sponse of normal lymphocytes to PHA. This are hyporesponsive to stimulation by phytoimmunosuppressive activity parallels in hemagglutinin (PHA) in vitro 125, 26, 281. degree and duration the severity of the This has been taken as evidence of a non- clinical course and appears to be localized specific depression of T-cell reactivity which in a low molecular weight ( ‘0.1) between bum and control cell ratios in either pooled or autologous serum. Examined individually, however, (Tables 4 and 5) it can be seen that while the cells of 4 of 8 controls had a wash index of 2 or greater in autologous serum (and hence were at least twice as reactive after six washes than after one wash), only 1 of 21 burn patients was as reactive. This difference was significant (P < 0.025) by x2 analysis. The differences in pooled serum (4 of 8 controls versus 7 of 21 patients) were not significant (P > 0.5). DISCUSSION
Recent evidence for post-traumatic lymphocyte hyporeactivity, especially to stimuli which require the participation of T-cells
CONSTANTIAN:
BURN
PATIENT
TABLE
89
LYMPHOCYTES
4
RESPONSE OF CELLS WASHED SIX TIMES TO PHA IN A~TOLOGO~S SERUM (cpm f SE) Patients
Controls Experiment No. 8 8 14 14 14 15 15 16 16 17 17 17 18 18 21 22 22 22 23 23 Mean f. SE
No PHA
295 2
PHA (l/200)
9
305 “_ 60 156 2
55
7,169 -c
633
13,952 rc_2,455 1,905 +
395
401 2 105
38,224 2 1,860
695 k 219
4,198 -+ 1,165
220 k 40 1,708 2 573
48,466 a 1,356 6,310 2 2,866
1,118 2
29,836 r
36
612 2 208
590
18,753 2 6,805
(1, 19, 20, 24-26, 28) has been presumed to explain in part the prevalence of lethal infectious complications following burns and other injuries. While this may be the case, the origin and nature of the (lymphocyte) hyporesponsiveness has not been established. Data from a number of laboratories have indicated that the serum of traumatized patients contain immunosuppressive activity which can inhibit antibodydependent cell-mediated cytolysis [30] and the response of peripheral blood lymphocytes to allogeneic cells [20], to Moniliu antigen [24], and to PHA [5, 71. This evidence has suggested that the observed host lymphocyte defects following trauma may be extrinsic and due to potentially reversible cell membrane changes brought about by a serum component. Our prior results indicate that in both surgically traumatized [7] and burned patients [5] a majority of the immunosuppressive serum activity is located in a low molecular
No PHA 138 ” 1.0 158 401 2 19 155 f 8 1,956 k 44 602 + 300 897 2 108 697 + 39 1,430 f 405 582 + 237 1,322 2 418 3,005 +- 184 629 k 267 2,051 2 568 660 k 346 2,033 2 103 589 r+ 51 907 2 80 1,291 4 96 923 5 103 1,198 -r- 252
PHA (l/200) 5,184 k 2,076 10,637 -+ 1,698 706 2 156 820 f 482 1,486 k 51 338 ? 42 1,037 f 37 158 4 44 4,956 2 570 3,681 + 177 3,745 2 947 5,448 c 864 588 2 174 846 -c 409 5,972 + 357 2,406 2 395 9,653 k 1,815 4,438 2 128 15,230 f 2,230 24,383 f 1,279 4,925 4 1,288
weight (
