JOURNAL OF SURGICAL RESEARCH25, 465-470 (1978)
Enzyme Activities in Experimental Skin Flaps 1 MICHAEL J.
IM,
PH.D., AND JOHN E.
HOOPES,
M.D.
Division of Plastic Surgery. The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205 Submitted for publication March 31, 1978 Seven enzymes participating in glucose catabolism were assayed under optimal assay conditions in the epidermis, dermis (papillary and reticular), and the panniculus carnosus at various regions of the pedicle skin flaps of guinea pigs. The skin, excluding the panniculus carnosus, exhibited minor alterations in enzyme activities in the proximal portions, considerable increases in glycolytic enzyme activities in the midregions, and progressive inactivation of all enzymes assayed in the distal end of the flaps. The panniculus carnosus displayed little alteration in glycolytic enzyme activities but a marked increase in glucose-6-phosphate dehydrogenase activity throughout the flaps.
INTRODUCTION
1, 2, and 3 days following flap elevation. The skin samples of the proximal portion Skin flap metabolism has been assessed were obtained from the midline of the back by means of measurement of pH, pO2, and at a distance of 1 cm from the pedicle of the pCO2 [1-3, 6, 7]. These investigations flap, the midregion was 4.5 cm, and the have substantiated ischemia and have, indistal biopsy was 8 cm from the pedicle. directly, revealed increased anaerobic Tissue blocks were frozen immediately in metabolism. Yet, there is no critical inliquid nitrogen. Preparation of lyophilized formation available for accurate interpretatissue sections and microdissection of tion of flap tissue metabolism. Microcutaneous organs were performed as deanalytical assays of a series of enzymes in scribed previously [5]. isolated samples of cutaneous organs would Seven of the key enzymes representing contribute to an understanding of the local the metabolic pathways of glucose were tissue metabolic alterations occurring assayed fluorometrically under optimal within skin flaps and would serve as a baseassay conditions [4]: hexokinase, phosline for further investigations concerning the phofructokinase (PFK), pyruvate kinase, "delay" phenomena. lactate dehydrogenase (LDH), isocitrate dehydrogenase (ICDH), malate dehydroMATERIAL AND METHODS genase (MDH), and glucose-6,phosphate Fifteen Hartley male guinea pigs weigh- dehydrogenase (G6PDH). Enzyme acing between 500 and 600 g were used in this tivities were assayed in 10/xl of the reagent investigation. Consistency of cutaneous mixture, utilizing 1- to 5-/xg samples of necrosis in the distal end of the pedicle flaps epidermis, papillary dermis, reticular of guinea pigs was determined [5]. Pedicle dermis, and panniculus carnosus. flaps, 3.5 x 9 cm, were elevated on the RESULTS backs of guinea pigs as described previously. Table 1 summarizes the distribution of Skin biopsies, 1 x 1 cm, were obtained seven enzyme activities in normal back from three different regions of the flaps skin. The present data demonstrate no reunder light ether anesthesia at intervals of gional variation in enzyme activities within i Supported by Grant AM 18732 of the National Institutes of Health.
normal back skin. However, a marked difference in the levels of enzyme activities
465
0022-4804/78/0255-0465501.00/0 Copyright© 1978by AcademicPress, Inc. All rightsof reproductionin any formreserved.
466
JOURNAL OF SURGICAL RESEARCH: VOL. 25, NO. 5, NOVEMBER 1978 TABLE 1 QUANTITATIVE ENZYME HISTOCHEMISTRY OF NORMAL BACK SKIN OF GUINEA PIGS a
Region of back skin Enzyme
Stratum of skin
Cephalic
Middle
Caudal
HK b
Epidermis Dermis: papillary reticular Panniculus carnosus
0.26 0.097 0.015 0.088
± ± _ ±
0.06 0.020 0.006 0.020
0.25 0.094 0.015 0.088
± 0.06 ± 0.029 ± 0.005 --_ 0.009
0.24 0.077 0.017 0.094
± ± ± ±
0.05 0.015 0.006 0.030
PFK
Epidermis Dermis: papillary reticular Panniculus carnosus
0.90 0.43 0.028 3.98
± 0.05 ± 0.09 ± 0.005 ±0.59
0.93 0.38 0.024 4.42
± 0.26 ± 0.06 ± 0.005 ±0.74
0.83 0.31 0.031 5.47
± _+ ± ±
0.26 0.06 0.004 3.15
PK
Epidermis Dermis: papillary reticular Panniculus carnosus
12.8 2.8 1.3 28.9
± ± ± ±
2.1 1.2 0.7 4.2
12.5 2.4 1.1 23.7
± ± ~ ±
1.4 1.3 0.3 5.7
13.8 4.4 1.6 32.4
± 2.0 _+ 0.8 _+ 0.2 _+ 11.6
LDH
Epidermis Dermis: papillary reticular Panniculus carnosus
15.4 7.4 1.6 33.0
± 2.3 ± 1.2 ± 0.4 _+ 2.6
14.5 6.0 1.6 34.5
+_ 4.1 ± 0.9 ± 0.9 + 2.0
16.3 6.0 1.8 38.9
_+ _+ _+ _+
9.2 1.4 0.6 5.6
G6PDH
Epidermis Dermis: papillary reticular Panniculus carnosus
0.57 0.27 0.085 0.034
±0.10 _+ 0.05 ± 0.028 ± 0.006
0.57 0.24 0.096 0.050
±0.05 ± 0.05 ± 0.025 ± 0.016
0.61 0.21 0.095 0.056
_+ -+ ± ±
0.12 0.06 0.034 0.019
ICDH
Epidermis Dermis: papillary reticular Panniculus carnosus
1.09 0.63 0.15 1.39
±0.11 _+ 0.06 ±0.03 ± 0.20
1.16 0.55 0.14 1.59
___0.13 ±0.11 ± 0.02 ± 0.65
1.21 0.57 0.18 1.97
_+ ± ± ±
0.11 0.10 0.06 0.33
MDH
Epidermis Dermis: papillary reticular Panniculus carnosus
± ± ± -+
2.8 3.4 0.3 9.8
24.0 14.7 3.1 23.5
± ± ± ±
3.6 2.4 0.7 3.8
24.1 10.9 2.9 25.9
± ± ± ±
3.6 1.0 0.7 3.9
25.8 13.7 3.5 26.6
a Enzyme activities are expressed as moles per hour per kilogram tissue dry weight _SD. Each figure is a total mean of 15 to 30 determinations obtained from three to six different animals. b Abbreviations used: HK, hexokinase; PFK, phosphofructokinase; PK, pyruvate kinase; LDH, lactate dehydrogenase; G6PDH, glucose-6-phosphate dehydrogenase; ICDH, isocitrate dehydrogenase; MDH, malate dehydrogenase. existed between epidermis, dermis, and panniculus carnosus. Glycolytic enzyme activities were greater in the panniculus carnosus than the overlying skin, whereas enzyme activity representing the pentose phosphate pathway in the subcutaneous muscle was one-tenth that in the epidermis. The dermis exhibited greater enzyme activities in the superficial layers than in the deep layers. The enzymatic alterations in
the proximal, medial, and distal portions of skin flaps are demonstrated in the epidermis, the superficial and deep layers of dermis, and the panniculus carnosus (Figs.
1-4). Epidermis.
Figure ! illustrates the enzymatic alterations in the proximal, medial, and distal portions of the skin flaps. Relatively minor alterations in enzyme activities were found in the proximal por-
467
IM A N D HOOPES: E X P E R I M E N T A L S K I N FLAPS
ENZYME ACTIVITIES ( % NORMAL )
,,.o 200
// / /
Proximal
/ ~Jo"j "~, /, /
150-
Medi01
Distal
x
!
~:.
/
/ o/
\
d'o I.X . . . . .
X-
/
I00-
X
50
0
,
.
1
2
,
3
0
DAYS
AFTER
,
,
J
I
2
5
0
I
,
,
I
2
3
FLAP ELEVATION
FIG. 1. Alterations in epidermal enzyme activities in the proximal, medial, and distal portions of the skin flaps. Each point was derived from a total mean of 15 to 20 determinations from three to four different skin samples. (O 0 ) hexokinase; ( 0 - - - 0 ) PFK; (O - O) pyruvate kinase; (O - - - O) LDH; (A L~) G6PD; (x -x) ICDH; ( x - - ×) MDH.
tion of the flaps. The epidermis of the midregions exhibited approximately a twofold increase in the activities of glycolytic enzymes and minor alterations in G6PDH, ICDH, and MDH activities during the 3 days following operation. Epidermal enzyme activities in the distal end of the flaps decreased progressively following flap elevation; a 60 to 90% decrease was found in all the enzyme activities by Day 3. Papillary dermis. No dramatic alterations in enzyme activities were found in the papillary dermis of the proximal portion of the flaps (Fig. 2). A slight increase in the glycolytic enzymes was noted in the midregion of the flaps: a 2.5-fold increase in
pyruvate kinase and an approximately 50% increase in hexokinase and lactate dehydrogenase activity by Day 3 postoperatively. PFK activity was in the normal level. The papillary dermis in the distal end of skin flaps exhibited decreases in PFK, pyruvate kinase, ICDH, and MDH activities. Relatively stable levels were found in hexokinase, lactate d e h y d r o g e n a s e , and G6PDH activities. Reticular dermis. Enzymatic alterations in the reticular dermis are illustrated in Fig. 3. No significant alterations in enzyme activities were demonstrated in the proximal portion. The reticular dermis of the midregion exhibited a threefold increase in
468
J O U R N A L O F S U R G I C A L R E S E A R C H : VOL. 25, NO. 5, N O V E M B E R 1978
ENZYME ACTIVITIES (% NORMAL) 250
i 200Proximal
150-
Distal
I00-
\
-,;\.¢-.~
\_
',.M,,//
. O . . .
°
50
0
I
I
I
I
2
3
0
I
I
I
I
2
3
0
I
I
I
I
2
3
DAYS AFTER FLAP ELEVATION FIG. 2. Alterations in e n z y m e activities in the papillary dermis of the skin flaps. L e g e n d is the same as in Fig. 1. ENZYME ACTIVITIES ( % NORMAL)
Distal
Mediol
Proximal 300"
/5"
200-
/,/" / / I I
[I
\
i
\k
x
I
//ix
x
~0------\\-.-~ //
C~'~
IO0-
r~ /
X.
~
\
I ~ ,
Nx
:]
0
I
I
I
2
I
'3
I
f
r
'0 I 2 '3 0 DAYS AFTER FLAP ELEVATIO~N
I
I
I
2
i
FIG. 3. Alterations in e n z y m e activities in the reticular dermis of the skin flaps. L e g e n d is the s a m e as in Fig. l.
469
IM A N D HOOPES: E X P E R I M E N T A L S K I N FLAPS
ENZYME ACTIVITIES ( % NORMAL) Medial
Proximal
]520
Distal
400
300
200-
//
\,
/__/2o
I00
0
I
[
I
I
2
3
0
I
~
I
I
2
3
O
I
2
3
DAYS AFTER FLAP ELEVATION FIG. 4. Alterations in enzyme activities in the panniculus carnosus of the skin flaps. Legend is the same as in Fig. l,
hexokinase and PFK and a twofold increase in pyruvate kinase and L D H activity by Day 3 postoperatively. A small magnitude of alterations in G6PDH, ICDH, and MDH activities was observed in the midregion. The reticular dermis of the distal end of flaps demonstrated an approximately 50% decrease in enzyme activities with the exception of hexokinase and L D H activity. Panniculus carnosus. Enzymatic reactions of the subcutaneous muscle to the flap elevation were different from those in the overlying skin (Fig. 4). The panniculus carnosus in the proximal portions of the flaps exhibited a marked increase in G6PDH activity and slight increases in glycolytic enzyme activities during the 3 days following flap elevation. In the midregion the muscle displayed a fivefold increase in G6PDH and a threefold increase in hexokinase activity by Day 3. Other
enzymes assayed exhibited a 50% increase in the midregions. The skin samples obtained from the distal end of the flaps represented tissue destined for necrosis [5]. However, the panniculus carnosus in the distal portion exhibited a marked increase in G6PDH and hexokinase activities and minor alterations in other enzymes assayed. DISCUSSION
The present data demonstrate different patterns of enzymatic alterations between the various regions and strata of skin flaps. In general, the epidermis and dermis exhibit normal levels of enzyme activities in the proximal portion of the flaps, increased enzyme activities in the midregion, and decreased activities in the distal end of the flaps. Minor alterations in enzyme activities in the proximal portion would be expected.
470
JOURNAL OF SURGICAL RESEARCH: VOL. 25, NO. 5, NOVEMBER 1978
The skin of the midregion, destined for dermis (Figs. 3 and 4). Skin necrosis probasurvival, demonstrates an example of bly is due to the inactivation of key enzymes metabolic adaptation of tissue to an altered in the enzyme machinery, together with environment. Although this flap tissue may limitation of glucose supply in the distal be partially ischemic [6] and low in pO2 extremity of skin flaps [5]. [2], abundant glucose content has been demonstrated in that portion of the flaps REFERENCES [5]. Increased enzyme activities, especially in epidermis, may reflect an enhanced 1. Glinz, W., and Clodius, L. Measurement of tissue pH for predicting viability in pedicle flaps: utilization of glucose through the glycolytic Experimental studies in pigs. Brit. J. Plast. Surg. pathway in the surviving flap tissues. A 25: 111, 1972. specific feature of skin metabolism--in2. Guthrie, R. H., Goulian, D., and Cucin, R. L. tense glyolytic activity in the presence of Predicting the extent of viability in flaps by measoxygen--is amplified by ischemia and urement of gas tensions using a mass spectrometer. Plast. Reconstr. Surg. 50: 385, 1972. possibly is responsible for the survival of 3. Hayes, J. E., Robinson, D. W., and Masters, this portion of the flap. F. W. A simple inexpensive method of evaluating The distal end of the flaps displays a circulation in pedicled tissues. Plast. Reconstr. progressive decrease with time in all Surg. 42: 141, 1968. enzyme activities in epidermis following flap 4. Ira, M. J., and Hoopes, J. E. Enzyme activities in the repairing epithelium during wound healing. elevation. Dermal tissues from the distal J. Surg. Res. 10: 173, 1970. end of the flap also exhibit decreases in 5. Ira, M. J., and Hoopes, J. E. Distribution of enzyme activities but of lesser magnitude glucose in the pedicle skin flaps of guinea pigs. than in epithelial tissues. Epidermal enzyme J. Surg. Res. 25: 269, 1978. assays can be utilized to predict potential 6. Myers, M. D., Cherry, G., and Milton, S. Tissue gas levels as an index of the adequacy of cirsurvival or necrosis of flap tissue. Hexoculation: The relation between ischemia and the kinase and LDH, participating in the initial development of collateral circulation. Surgery 71: and final step of glycolysis, are relatively 15, 1972. stable in dermis; PFK and pyruvate kinase 7. Toomey, J. M., and Wilson, W. R. Mass spectro(of the intermediate steps) exhibit conmetric observations of skin flap physiology. Laryngoscope 83: 559, 1973. siderable decreases in their activities in the
Enzyme Activities in Experimental Skin Flaps 1 MICHAEL J.
IM,
PH.D., AND JOHN E.
HOOPES,
M.D.
Division of Plastic Surgery. The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205 Submitted for publication March 31, 1978 Seven enzymes participating in glucose catabolism were assayed under optimal assay conditions in the epidermis, dermis (papillary and reticular), and the panniculus carnosus at various regions of the pedicle skin flaps of guinea pigs. The skin, excluding the panniculus carnosus, exhibited minor alterations in enzyme activities in the proximal portions, considerable increases in glycolytic enzyme activities in the midregions, and progressive inactivation of all enzymes assayed in the distal end of the flaps. The panniculus carnosus displayed little alteration in glycolytic enzyme activities but a marked increase in glucose-6-phosphate dehydrogenase activity throughout the flaps.
INTRODUCTION
1, 2, and 3 days following flap elevation. The skin samples of the proximal portion Skin flap metabolism has been assessed were obtained from the midline of the back by means of measurement of pH, pO2, and at a distance of 1 cm from the pedicle of the pCO2 [1-3, 6, 7]. These investigations flap, the midregion was 4.5 cm, and the have substantiated ischemia and have, indistal biopsy was 8 cm from the pedicle. directly, revealed increased anaerobic Tissue blocks were frozen immediately in metabolism. Yet, there is no critical inliquid nitrogen. Preparation of lyophilized formation available for accurate interpretatissue sections and microdissection of tion of flap tissue metabolism. Microcutaneous organs were performed as deanalytical assays of a series of enzymes in scribed previously [5]. isolated samples of cutaneous organs would Seven of the key enzymes representing contribute to an understanding of the local the metabolic pathways of glucose were tissue metabolic alterations occurring assayed fluorometrically under optimal within skin flaps and would serve as a baseassay conditions [4]: hexokinase, phosline for further investigations concerning the phofructokinase (PFK), pyruvate kinase, "delay" phenomena. lactate dehydrogenase (LDH), isocitrate dehydrogenase (ICDH), malate dehydroMATERIAL AND METHODS genase (MDH), and glucose-6,phosphate Fifteen Hartley male guinea pigs weigh- dehydrogenase (G6PDH). Enzyme acing between 500 and 600 g were used in this tivities were assayed in 10/xl of the reagent investigation. Consistency of cutaneous mixture, utilizing 1- to 5-/xg samples of necrosis in the distal end of the pedicle flaps epidermis, papillary dermis, reticular of guinea pigs was determined [5]. Pedicle dermis, and panniculus carnosus. flaps, 3.5 x 9 cm, were elevated on the RESULTS backs of guinea pigs as described previously. Table 1 summarizes the distribution of Skin biopsies, 1 x 1 cm, were obtained seven enzyme activities in normal back from three different regions of the flaps skin. The present data demonstrate no reunder light ether anesthesia at intervals of gional variation in enzyme activities within i Supported by Grant AM 18732 of the National Institutes of Health.
normal back skin. However, a marked difference in the levels of enzyme activities
465
0022-4804/78/0255-0465501.00/0 Copyright© 1978by AcademicPress, Inc. All rightsof reproductionin any formreserved.
466
JOURNAL OF SURGICAL RESEARCH: VOL. 25, NO. 5, NOVEMBER 1978 TABLE 1 QUANTITATIVE ENZYME HISTOCHEMISTRY OF NORMAL BACK SKIN OF GUINEA PIGS a
Region of back skin Enzyme
Stratum of skin
Cephalic
Middle
Caudal
HK b
Epidermis Dermis: papillary reticular Panniculus carnosus
0.26 0.097 0.015 0.088
± ± _ ±
0.06 0.020 0.006 0.020
0.25 0.094 0.015 0.088
± 0.06 ± 0.029 ± 0.005 --_ 0.009
0.24 0.077 0.017 0.094
± ± ± ±
0.05 0.015 0.006 0.030
PFK
Epidermis Dermis: papillary reticular Panniculus carnosus
0.90 0.43 0.028 3.98
± 0.05 ± 0.09 ± 0.005 ±0.59
0.93 0.38 0.024 4.42
± 0.26 ± 0.06 ± 0.005 ±0.74
0.83 0.31 0.031 5.47
± _+ ± ±
0.26 0.06 0.004 3.15
PK
Epidermis Dermis: papillary reticular Panniculus carnosus
12.8 2.8 1.3 28.9
± ± ± ±
2.1 1.2 0.7 4.2
12.5 2.4 1.1 23.7
± ± ~ ±
1.4 1.3 0.3 5.7
13.8 4.4 1.6 32.4
± 2.0 _+ 0.8 _+ 0.2 _+ 11.6
LDH
Epidermis Dermis: papillary reticular Panniculus carnosus
15.4 7.4 1.6 33.0
± 2.3 ± 1.2 ± 0.4 _+ 2.6
14.5 6.0 1.6 34.5
+_ 4.1 ± 0.9 ± 0.9 + 2.0
16.3 6.0 1.8 38.9
_+ _+ _+ _+
9.2 1.4 0.6 5.6
G6PDH
Epidermis Dermis: papillary reticular Panniculus carnosus
0.57 0.27 0.085 0.034
±0.10 _+ 0.05 ± 0.028 ± 0.006
0.57 0.24 0.096 0.050
±0.05 ± 0.05 ± 0.025 ± 0.016
0.61 0.21 0.095 0.056
_+ -+ ± ±
0.12 0.06 0.034 0.019
ICDH
Epidermis Dermis: papillary reticular Panniculus carnosus
1.09 0.63 0.15 1.39
±0.11 _+ 0.06 ±0.03 ± 0.20
1.16 0.55 0.14 1.59
___0.13 ±0.11 ± 0.02 ± 0.65
1.21 0.57 0.18 1.97
_+ ± ± ±
0.11 0.10 0.06 0.33
MDH
Epidermis Dermis: papillary reticular Panniculus carnosus
± ± ± -+
2.8 3.4 0.3 9.8
24.0 14.7 3.1 23.5
± ± ± ±
3.6 2.4 0.7 3.8
24.1 10.9 2.9 25.9
± ± ± ±
3.6 1.0 0.7 3.9
25.8 13.7 3.5 26.6
a Enzyme activities are expressed as moles per hour per kilogram tissue dry weight _SD. Each figure is a total mean of 15 to 30 determinations obtained from three to six different animals. b Abbreviations used: HK, hexokinase; PFK, phosphofructokinase; PK, pyruvate kinase; LDH, lactate dehydrogenase; G6PDH, glucose-6-phosphate dehydrogenase; ICDH, isocitrate dehydrogenase; MDH, malate dehydrogenase. existed between epidermis, dermis, and panniculus carnosus. Glycolytic enzyme activities were greater in the panniculus carnosus than the overlying skin, whereas enzyme activity representing the pentose phosphate pathway in the subcutaneous muscle was one-tenth that in the epidermis. The dermis exhibited greater enzyme activities in the superficial layers than in the deep layers. The enzymatic alterations in
the proximal, medial, and distal portions of skin flaps are demonstrated in the epidermis, the superficial and deep layers of dermis, and the panniculus carnosus (Figs.
1-4). Epidermis.
Figure ! illustrates the enzymatic alterations in the proximal, medial, and distal portions of the skin flaps. Relatively minor alterations in enzyme activities were found in the proximal por-
467
IM A N D HOOPES: E X P E R I M E N T A L S K I N FLAPS
ENZYME ACTIVITIES ( % NORMAL )
,,.o 200
// / /
Proximal
/ ~Jo"j "~, /, /
150-
Medi01
Distal
x
!
~:.
/
/ o/
\
d'o I.X . . . . .
X-
/
I00-
X
50
0
,
.
1
2
,
3
0
DAYS
AFTER
,
,
J
I
2
5
0
I
,
,
I
2
3
FLAP ELEVATION
FIG. 1. Alterations in epidermal enzyme activities in the proximal, medial, and distal portions of the skin flaps. Each point was derived from a total mean of 15 to 20 determinations from three to four different skin samples. (O 0 ) hexokinase; ( 0 - - - 0 ) PFK; (O - O) pyruvate kinase; (O - - - O) LDH; (A L~) G6PD; (x -x) ICDH; ( x - - ×) MDH.
tion of the flaps. The epidermis of the midregions exhibited approximately a twofold increase in the activities of glycolytic enzymes and minor alterations in G6PDH, ICDH, and MDH activities during the 3 days following operation. Epidermal enzyme activities in the distal end of the flaps decreased progressively following flap elevation; a 60 to 90% decrease was found in all the enzyme activities by Day 3. Papillary dermis. No dramatic alterations in enzyme activities were found in the papillary dermis of the proximal portion of the flaps (Fig. 2). A slight increase in the glycolytic enzymes was noted in the midregion of the flaps: a 2.5-fold increase in
pyruvate kinase and an approximately 50% increase in hexokinase and lactate dehydrogenase activity by Day 3 postoperatively. PFK activity was in the normal level. The papillary dermis in the distal end of skin flaps exhibited decreases in PFK, pyruvate kinase, ICDH, and MDH activities. Relatively stable levels were found in hexokinase, lactate d e h y d r o g e n a s e , and G6PDH activities. Reticular dermis. Enzymatic alterations in the reticular dermis are illustrated in Fig. 3. No significant alterations in enzyme activities were demonstrated in the proximal portion. The reticular dermis of the midregion exhibited a threefold increase in
468
J O U R N A L O F S U R G I C A L R E S E A R C H : VOL. 25, NO. 5, N O V E M B E R 1978
ENZYME ACTIVITIES (% NORMAL) 250
i 200Proximal
150-
Distal
I00-
\
-,;\.¢-.~
\_
',.M,,//
. O . . .
°
50
0
I
I
I
I
2
3
0
I
I
I
I
2
3
0
I
I
I
I
2
3
DAYS AFTER FLAP ELEVATION FIG. 2. Alterations in e n z y m e activities in the papillary dermis of the skin flaps. L e g e n d is the same as in Fig. 1. ENZYME ACTIVITIES ( % NORMAL)
Distal
Mediol
Proximal 300"
/5"
200-
/,/" / / I I
[I
\
i
\k
x
I
//ix
x
~0------\\-.-~ //
C~'~
IO0-
r~ /
X.
~
\
I ~ ,
Nx
:]
0
I
I
I
2
I
'3
I
f
r
'0 I 2 '3 0 DAYS AFTER FLAP ELEVATIO~N
I
I
I
2
i
FIG. 3. Alterations in e n z y m e activities in the reticular dermis of the skin flaps. L e g e n d is the s a m e as in Fig. l.
469
IM A N D HOOPES: E X P E R I M E N T A L S K I N FLAPS
ENZYME ACTIVITIES ( % NORMAL) Medial
Proximal
]520
Distal
400
300
200-
//
\,
/__/2o
I00
0
I
[
I
I
2
3
0
I
~
I
I
2
3
O
I
2
3
DAYS AFTER FLAP ELEVATION FIG. 4. Alterations in enzyme activities in the panniculus carnosus of the skin flaps. Legend is the same as in Fig. l,
hexokinase and PFK and a twofold increase in pyruvate kinase and L D H activity by Day 3 postoperatively. A small magnitude of alterations in G6PDH, ICDH, and MDH activities was observed in the midregion. The reticular dermis of the distal end of flaps demonstrated an approximately 50% decrease in enzyme activities with the exception of hexokinase and L D H activity. Panniculus carnosus. Enzymatic reactions of the subcutaneous muscle to the flap elevation were different from those in the overlying skin (Fig. 4). The panniculus carnosus in the proximal portions of the flaps exhibited a marked increase in G6PDH activity and slight increases in glycolytic enzyme activities during the 3 days following flap elevation. In the midregion the muscle displayed a fivefold increase in G6PDH and a threefold increase in hexokinase activity by Day 3. Other
enzymes assayed exhibited a 50% increase in the midregions. The skin samples obtained from the distal end of the flaps represented tissue destined for necrosis [5]. However, the panniculus carnosus in the distal portion exhibited a marked increase in G6PDH and hexokinase activities and minor alterations in other enzymes assayed. DISCUSSION
The present data demonstrate different patterns of enzymatic alterations between the various regions and strata of skin flaps. In general, the epidermis and dermis exhibit normal levels of enzyme activities in the proximal portion of the flaps, increased enzyme activities in the midregion, and decreased activities in the distal end of the flaps. Minor alterations in enzyme activities in the proximal portion would be expected.
470
JOURNAL OF SURGICAL RESEARCH: VOL. 25, NO. 5, NOVEMBER 1978
The skin of the midregion, destined for dermis (Figs. 3 and 4). Skin necrosis probasurvival, demonstrates an example of bly is due to the inactivation of key enzymes metabolic adaptation of tissue to an altered in the enzyme machinery, together with environment. Although this flap tissue may limitation of glucose supply in the distal be partially ischemic [6] and low in pO2 extremity of skin flaps [5]. [2], abundant glucose content has been demonstrated in that portion of the flaps REFERENCES [5]. Increased enzyme activities, especially in epidermis, may reflect an enhanced 1. Glinz, W., and Clodius, L. Measurement of tissue pH for predicting viability in pedicle flaps: utilization of glucose through the glycolytic Experimental studies in pigs. Brit. J. Plast. Surg. pathway in the surviving flap tissues. A 25: 111, 1972. specific feature of skin metabolism--in2. Guthrie, R. H., Goulian, D., and Cucin, R. L. tense glyolytic activity in the presence of Predicting the extent of viability in flaps by measoxygen--is amplified by ischemia and urement of gas tensions using a mass spectrometer. Plast. Reconstr. Surg. 50: 385, 1972. possibly is responsible for the survival of 3. Hayes, J. E., Robinson, D. W., and Masters, this portion of the flap. F. W. A simple inexpensive method of evaluating The distal end of the flaps displays a circulation in pedicled tissues. Plast. Reconstr. progressive decrease with time in all Surg. 42: 141, 1968. enzyme activities in epidermis following flap 4. Ira, M. J., and Hoopes, J. E. Enzyme activities in the repairing epithelium during wound healing. elevation. Dermal tissues from the distal J. Surg. Res. 10: 173, 1970. end of the flap also exhibit decreases in 5. Ira, M. J., and Hoopes, J. E. Distribution of enzyme activities but of lesser magnitude glucose in the pedicle skin flaps of guinea pigs. than in epithelial tissues. Epidermal enzyme J. Surg. Res. 25: 269, 1978. assays can be utilized to predict potential 6. Myers, M. D., Cherry, G., and Milton, S. Tissue gas levels as an index of the adequacy of cirsurvival or necrosis of flap tissue. Hexoculation: The relation between ischemia and the kinase and LDH, participating in the initial development of collateral circulation. Surgery 71: and final step of glycolysis, are relatively 15, 1972. stable in dermis; PFK and pyruvate kinase 7. Toomey, J. M., and Wilson, W. R. Mass spectro(of the intermediate steps) exhibit conmetric observations of skin flap physiology. Laryngoscope 83: 559, 1973. siderable decreases in their activities in the
