close

Вход

Забыли?

вход по аккаунту

?

Histology and cytochemistry of human skin. VII. The distribution of succinic dehydrogenase activity

код для вставкиСкачать
HISTOLOGY AND CYTOCHE,I\llSTRY O F HUMAN SKIN
VII.
THE DISTRIBUTION O F SUCCINIC DEHYDROGENASE
-4CTIVITY
WILLIAM MOKTAGNA AND VICTOR FORMISANO
Arnold Biological Laboratory, Brown University,
Providence, Blbode Island
ELEVEN FIGURES
INTRODUCTION
The epidermis and its appendages contain relatively large
amounts of demonstrable succinic dehydrogenase activity,
although not all of the appendages have equal amounts of
it. The distribution of succinic dehydrogenase in skin suggests that the enzyme is involved in both metabolic and
proliferative functions. I n the matrix of growing hair follicles
and in the basal layer of the epidermis, the enzyme must
be concerned mainly with proliferative activity, whereas that
in the outer sheath of hair follicles and in the eccrine sweat
glands, both of which a r e niitotically inert, must be related
to metabolic functions.
This paper reports the localization of succinic dehydrogeiiase activity in fresh pieces of normal human skin. Previously, only Rogers ('53) made some observations of the
distribution of the enzgmc in human skin. Ellis et al. ('51),
and Padykula ( '52) refer briefly to succinic dehydrogenase
in the skin of laboratory animals, and Rogers ('53) and Forrnisano and JIonttigna ('2)
made thorough studies of the enzyme in the skin of the shclep and the guinea pig, respectively.
'This work was nipportcd in p a r t by a g r a n t f r o m t h e Vnited States Public
Health Service, RG 2125 C i .
65
66
WILLIAM MONTAGNA A N D VICTOR F O R M I S A N O
MATERIALS ,4KD RlETIiODS
Sixteen pieces of skin were used in this study. The
specimens were all removed without anesthesia with a motordriven rotary biopsy punch 5 mm (Urbacli and Shelley, '51 ).
Five pieces of axilla, three pieces of scalp, three pieces of
chest, 4 pieces of palm, and one of back were rcniorc3d from
normal male and female subjects ranging from 24 to 34 pears
of age. Each piece of skin was immediately pinned, epidermis
down, on a paraffin l)lock, and as much f a t as possible way
trimmed away from the hypodermis. The operation, which
was never proloiigcd beyond two minutes, was performed
under a dissecting microscope, and the tissue was kept bathed
with warni physiological saline. The bulbs of hair follicles
and the coils of sweat glands were partially freed from the
surrounding connective tissue and fat (Formisaiio and Montagna, '54). Each piece w a s cut into two, and both fragments
were immersed in the incuhatiiig mixture.
The incubating mixture was prepared according to Pad).kula's ('5%) inodificntion of the method of Seligman and
Kutenburg ( '51), using neotetrazolium ( N T ) as the hydrogen
acceptor. CYalciunichloride and aluminum chloride mere acldcd
to the mixture since the ions of these salts a r e said to activatc
the succinic ciehydrogenase system (Padykula, ' 3 2 ) . 13cforc.
the solution was uscd it w a s boiled, the flask \vas st opperecl and
allowed to cool to 37°C. When the pieces of tissue were placed
in the solution the flask n7as closed tightly with a rubher stopper fitted with two glass tubes, one of which was attached
to a nitrogen tank. The tissues ivere incubated for two hours
at 37°C. with nitrogen gas lmbhling slowly through tlic mixture.
After incubation the tissues were placed in 20% formalin
and stored i n the refrigerator for 24 hoiirs. The tissues were
then cut in sections 15 p thick with the freezinq microtome;
the sections were hardened in 10% formalin and then mounted
in glycerin.
SUCCINIC DEHYDROGENASE I N H U M A N S K I N
67
OBSERVATIONS
The epidermis. I n the epidermis of the scalp, the axilla,
the back and the chest, the strongest enzyme reaction, indicated by the presence of intracellular, purple diformazan crystals, is found in the basal cells. The lower two or three
layers of the stratum spinosum contain variable amounts
of reactive crystals but the reaction fades gradually upwards
and disappears completely in the cells just below the stratum
granulosum (fig. 1). The rest of the epidermis is unreactive.
The cytoplasmic radicles on the lower surface of the basal
cells arc strongly reactive. I n the thick epidermis of the
palms, tlie reaction is soniewhat stronger and more extensive but it also disappears below the stratum granulosum.
14s in the other epidermis, tlie reaction is strongest in the basal
layer. Since the reaction in the epidermis is much stronger in
some preparations than in others, the penetration of the substrate and dye may have been uneven. Penetration seems to
take place through the dermis, the stratum corneum serving as
a barrier.
I n the intraepidermal portion of tlie ducts of the eccrine
sweat glands, only the lower segment is reactive. The periductual and the basal cells of the duct show formazan granules,
but the luniinal cells arc’ unreactive. The weak reaction in the
“eccrine sweat duct unit” of Lobitz et al. (54) may be due
t o a lack of proper penetration by the substrate and dye
through the epidermis.
The pilosebnccous S ! ~ S ~ C I W . The distribution of the formazan granules in the wall of the pilosebaceous canal is
similar to but stronger than that in the epidermis
I n growing hair follicles, the entire outer sheath is rich in
enzyme activity (fig.2). The reaction is particularly brilliant
around the lower third of the follicle, the cells of which are
riddled with vacuoles which contain glycogen and mucopolysaccharide. The cells in the periphery of the outer sheath are
always more strongly reactive than the others (fig. 3). I n the
upper part of the hair bulb, the non-keratinized portion of the
inner sheath and of the cortex show moderate enzymatic
GS
IVILLIAM M O N TAGKA A N D VICTOR FORMISANO
reaction. Thc n i a l ~ - i sor
, blie lowei- bulb, which is easily recognized from the upper bulb by the absence of pigment, is
intensely reactive. This is not shown well by figure 2. The
fibroblasts ;md cndotlielial cells in the dcriiial papilla, and the
fihroblasts in thc coiinectivc tissue sheath contain rcactive
granules.
Thc follicles of club hairs contain much less succinic dehydrogenase activity. The entire outer sheath and the epidermal sac arc only nioderatcly rcactive. The cord of cells
hetween tlic epidermal sac and the hall-like dermal papilla,
often referred to a s the “hair germ,” has only some enzyme
activity. The deimiil papilla also sliows reduced enzyme
action.
I n the sebaceous glmids, the cells of the duct system, and
the indiff’erent sebaceous cells usually found a t the periphery
of the glandular acini show strong reaction (fig. 4). The
f ormazan crysials gradually diminish a s the cells accuniulate
lipid globules, and they disappear in niatuwe sehaceous cells.
The s.ebum is c o l o r d pink but this may he due to a diffusion
of tlic formazan crystals which a r e solulnle in fats.
Siucnt gltrntls. The eccrine sweat glands and their ducts
a r e tlic most reactive eleineiits in huniari skin. In tissues which
liave heen teased rat her. than sectioned, entire s w e a t glands
can he seen against the unstained background of the dermis
(fig. 3 ) . These ~ ~ r e p a i . a t i o iai sr e useful f o r the demonstration
of thcl gross niorpholog:.y of the glands, and the glands can
he traced from the epidermis d o ~ wto the secretory knot.
These preparations show that the eccririe sweat glands are
considerably longer than is usually aclinowledged.
I n the straight clscretory duct, which extends from the
epidermis to the secretory coil, the cytoplasm of the basal
cells is laden with foriiiazan crystals. I n the luntinal cells,
rcactive graiiules are gatlimed around the nuclcus and a few
may he fourid in the cuticle, midway hctwecn the surface of
the cell and the lateral surface of the nucleus. The ducts with
a well-dcvclopcd cuticle havc fewer reactive granules in the
luniinal cells.
ST'CCIKIC
DEHYDROGENASE I N HVlRIhN S K I N
69
The coiled p a r t of tlie gland is coinposed of equal parts of
duct and secretory tuhule. The coiled duct usually, but not
always, has a poorly developed cuticle, and a segment of the
duct seems to be the ti.ansition zone (,1Tontagna et al., '53).
Both the hasal illid luniinnl cells of tlie coiled and transition
part of tlic duct show very strong exizyrne activity, although
the surface of the luniiiial cells is less reactive (figs. 6, 7 ) .
The secretory coil of eccrine glands is the most intensely
reactive tissue in human skin. The reaction is occasionally
uneven, some cells showing greater. activity thaii others, but
as a rule, each cell is virtually 1;idcn with very dark purple
forinazari granules. The luniiiial part of the cells has fewer
granules than the hasal portion. The niyoepithelid cells seem
t o be unreactivc. I n the axilla a i d in the palm, the cccrine
glands a r e always iiiuch m o r e iiiteiisely c o l o r d than those
in the other skin studied. This seems to be a distinctive
feature of the glands in the pal111 aiid axilla.
I n contrast with the cccrine glands, tlie axillai-y a p o c ~ i n e
glands contain only small amouiits of succinic dehydrogeiiasc
(fig. 9 ) . The excretory ducts have a similar but weaker rcaction than the eccrine tlucts. 1x1 the secretoi-y cells, formaxan
granules a r e inversely proportional to the amount of secrctioxi material within the cells, regard1
of the size of thc
cells (figs. 10, 11). \Then the cells are full of secretion granules, formazaxi crystals a r e the least abundant and they ai*e
concentrated iii a region above the nucleus (fig.11). Formazan
granules a r e usually clustered around the nucleus or above
it. The basal part of the cells and the terminal cytoplasmic
border are usually free of i*eactive granules. The myoepithelial cells a r e consistently unveactive.
IlIisceZZcizzeous. The arrectorcs pilorum muscles contain
iiumerous reactive granules alligned orderly on the long axis
of the muscle fibers. The endothelid cells in the dermis a r e
always strongly reactive. The cytoplasni of fibroblasts has
a sparse granulation. In the mast cells, some forxiiazan granules may be formed between the mast granules, above the
nucleus.
70
WILLIAM MONTAGKA A N D VICTOR FORMISANO
DISCUSSION
This test cannot be performed adequately in fresh, frozen
sectioiis of skin. Enzyme activity in sections is minimal. When
wholc pieces of the saiiie skin a r e used, enzyme activity is
al)undant, provided that the f a t is removed from its underiide (cf. Formisaiio and Nontagiia, '54). Since it is not possible to remove all of the fat speedily enough so that the
remaining tissue is alive anti uiidaniaged when jt is placccl
in the incubating mixture, tlie penetration of the suhstr*ate
aiid dye through tlie tissue is not always optimal. Itefinenicrits are needed in this method hefoi*e perfect result.: a r e
o1)tained.
The abundaiit succinic dehydrogenase activity in the eccrine
+ w a t glands is in accord with the fact that succinic dehydi-ogenase f o r m s a link in the respiratory processes concerned
ivith the oxidation of lipids, carbohydrates, arid proteins
(Summer and Soinei.~,'47). The iich glycogen storage iii the
secretory tubule of sweat glands (Montagna et al., '52) can
bc exhausted temporarily hy prolonged stimulation (Shelley
arid AIesco~i, '52),and it should be interesting to know if the
distributioii of succiiiic dehydrogeiiase undergoes shifts cornparuhle t o those of the glycogc~ri. Finding succinic deliydrogenasc activity i n the ducts of sweat glands adds soine support
t o the already existing presumptive evidence that the eccrine
sivpat ducts have a physiologic function (Montagna ct al.,
'3).
The secretory activity of cccrine glands is practically incshaustible, but that of the apocrine glands is of very short
duration (Shelley, '51). One of the bases for thew differences
might be the great disparity in succinic dehydrogenase activity between these t\vo glands. Different segments of the
balm apocrine tubule oftcii show differences in the content
of the enzyme, depending upon thc ainourit of secretion granules within the cells. A similar situatioii exists in the sebaceous glands where the indiffcrent cells 211-e rich in enzyme
activity lmt the differentiated cc.11~a r e not.
SUCCIPI’IC IIEHYDROGENASE I N H I J M A N SKIN
71
The possihle significaiice of succiiiic dehydrogenase activity
in tissues of high mitotic activity has been discussed by
B’ormisaiio and Rlontagiia (’51)and will not be repeated here.
The distribution of the forniazari crystals in tissues is similar to that of the mitochondria. I n tlie eccrine sweat glands,
however, which :we particularly rich i n enzyme activity, the
number of mitochondria denionstruble with standard methods
is much smaller than that of the formazan crystals. I n apocrine cells, the correspondence is much closer. I n spite of
this discrepancy, it seems certain that the mitochondria arc’
the reactive foci of the enzyme a s stated before by other
authors (Padyliula, ’52 ; Shelton aiid Schneider, ’52 ; lllalaty
arid Bourne, ’53; Forrriisaiio arid Xontagna, ’54).
SUhfMhRV
1. Succinic dehytlrogenase activity is demonstrated more
successfully in small, x7hole pieces of sliiii than in fresh
frozen sections. I n order that the substrate and dye penctratc readily, it is necessary to remove as much fat from the
tcla subcutanea a s possible.
2. Succinic dehydrogenase activity is present in the basal
layer of the epidermis, in the outer sheath and bulb of hair
follicles and in m e a t glands. The reaction is most intense
in the eccrine sweat glands aiid iii their ducts. The apocriiie
glands show only moderate activity.
3. The distribution of the enzyme in skin strongly suggests
that the cnzyine is concerned with both metabolic and proliferative functions.
LITERATlTRE C I T E D
ELLIS,W. J., J. M. GILLESPIE , m n H. T d m r . E Y 1948 Biocheniical studics of
the wool root. Nature, 165: 545-548.
F O R X I S ~V.,
N OBNn
, W. MONTI O X L 1951 Succinic drliydrogenaxr activity in t h e
skin of tlie guinea pig. Anat. Rec., 1 W : 893-906.
LOBITZ,
W. C., JR., J. B. HOLYOKE4 ~ TqT.
n >fONTAGN4 1951- “ T h e epidermal
eccrine sweat duct unit.’ ’ A morpliologic and biologic entity. J. Invest.
Drrmatol., 5 2 : 157-158.
19.53 Historlicniistrv of succinic dehydroM 4 L A T Y , 1%.A , , ANI) G. 11. B O U R S E
genase. Nature, 77 I : 295-297.
72
W I L L I A M M O N T A G N A A N D VICTOR P O R M I S A N O
MONTAGNA,
W., H. B. CHASE BND TIT. C. LOBITZ,
JR. 19.72 Histology a d eytoclieinistry of human skin. 11. The distribution of glycogcn in the
epidermis, hair follicles, sebaceous glands aiid eceriiie sweat glauds.
Anat. Rec., 1 1 4 : 231-248.
PADYKULA,
H. A. 1952 The localization of succinic dehydrogennse i n tissne
sections of the r a t . Am. J. Anat., 9 1 : 107-146.
ROGERS,G. E. 1953 The localization of deliyclrogenase activity xird sulfhytlryl
groups i n wool arid hair follicles by the use of tetrazoliuni salts. Quart.
J. Micro. Sci., 94: 253-268.
SELIGMAN,
A. M., AND A. M. RUTENBURG1951 The histolocliemical dcnioiist r:ition of succinic dehydrogenase. Science, 115 : 31 7-320.
SHELLEY,
W. B. 1951 Apocrine sweat. J. Invest. Dermatol., 1 7 : 255.
SHELLEY,
W. B., AND If. MESCON 1952 Histochemical demonstration of secretory
activity i n human eccriiie sweat glancls. J. Invest. nermatol., IX:
289-301.
RHELTON, E., AND W. C. SCHNEIDER
1952 On the usefulness of tctrazoliuni
salts as histochemical indicators of dehydrogeriase activity. Anat. Rec.,
112: 61-82.
SUMNER,
J. B., AND G. F. SOMERS1947 Chemistry aiid Mcthods of Enzymes.
2nd ed. Academic Press, New York.
URBACH,
F.,AND W. B. SHELLEY1951 A rapid and simple method f o r obtaining
punch biopsies without anesthesia. J. Invest. Dermntol., 27 : 131-1 34.
PLATE 1
EXPLANATION O F FIGURES
1 Suecinic dehydrogenase activity i n the epidermis of the scalp. Formazaii
crystals concentrated in the basal layers of the stratum Malpighii. The
stratum granulosum (arrow) and the stratum corneuni are unreactive. X 550.
2
Enzyme reaction i n a hair follicle from the scalp. With the exception of the
dark cone above the dermal papilla ( P ) which is pigment, all otlicr dark
material represents enzyme activity. Poorly oriented f olliclc not slio\viiig
a n optimum amount of activity. X 250.
3
Formazan crystals i n the outer sheath of active hair follicle. Heaviest concentration i n the peripheral cells next t o the glassy membrane (arrow). X 500.
4 Distribution of the enzyme in a sebaceous gland from the back. The enzyme
is concentrated at the periphery of the aciiius and i n the duct. X 500.
SCCClNlC DEIIYDROUENABE I N HUMAN SKIN
WIIILUY MONTABXA A N D VICTOB FOBYISANO
73
PLATE 2
EXPLANATION OF FIGURES
5 Whole eccrine sweat gland from the axilla. X 60.
6 Eccrine sweat ducts from the palm. The heaviest concentration of the enzyme
is in the basal cells; the cuticle is nearly free of it. X 500.
7
Transition portion of the eccrine sweat duct. Reactive granules are abundant
up to the luminal border. From the palm. X 750.
8
Eccrine sweat gland; secretory tubule from the axilla. Extremely high eoncentration of reactive granules. X 750.
74
PLATE
SUCC.rN10 DEHYDROBENASE IN HUXAN BKIN
V I l J ~ I A MMONTAOIA AND VIDTOK XYIRMISANO
75
2
PLATE 3
EXPLANATION O F FIGUltES
9
Wliolc tubule of axillary a p o u i n e yl:md. Contrast with figure 5 . X GO.
10
Ahiiiitlant rrizyiiie activity
tion grmules. x 750.
11
Moilcrate iiiiniher of graiiulcs roiiceiitr atecl above the nucleus of apocriiic
cellz modrratrly laden with seeretioii grmiules. X 750.
iii
npocriiie cells which arc nearly free of secre-
76
PLATE
SUCCINIC DEHYDROBENASE IN HUJdAN SKIN
W I l r b I A I XONTAUBA AKD VICTOR. FORMIMNO
77
8
Документ
Категория
Без категории
Просмотров
1
Размер файла
1 142 Кб
Теги
histology, vii, distributions, succinic, cytochemistry, activity, dehydrogenase, skin, human
1/--страниц
Пожаловаться на содержимое документа