An experimental study of the effects of steroids on the osmotic fragility of red blood cells

60 Pages·02.489 MB·English

by  Abrahamson, Joseph

AN EXPERIMENTAL STUDY OP THE EFFECTS OF STEROIDS
ON THE OSMOTIC FRAGILITY OF RED BLOOD CELLS
A Thesis
Presented to
the Faculty of the Department of Physiology
School of Medicine
University of Southern California
In Partial Fulfillment
of the Requirements for the Degree
Master of Science
by
Joseph Abrahamson
September 1950
UMI Number: EP63578
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This thesis, written by
JOSEPH ABRAHAMSON
under the guidance of h..^.. Faculty Committee,
and approved by all its members, has been
presented to and accepted by the Council on
Graduate Study and Research in partial fulfill­
ment of the requirements for the degree of
MASTER OF SCIENCE
.....
rw - September 1950__
Faculty Committee
C . V
Chc^rhtan
...
TABLE OP CONTENTS
CHAPTER PAGE
I. THE PROBLEM........................... 1
Statement of the problem.................. .. 1
Importance of the study . . . . ........... 1
Preview of organization of thesis ......... 1
II. REVIEW OF THE LITERATURE ............ 3
The state of the problem when this project
was b e g u n ................................ 3
III. EXPERIMENTAL PROCEDURE . . . . .............. 7
Method used for this investigation........ 7
IV. DISCUSSION.................. 15
Procedure....................... 15
The red blood cell membranes in general
and this problem in particular........... 15
V. EVALUATION OF RESULTS AND COMMENTS.......... 28 *
VI. SUMMARY AND CONCLUSIONS..................... 32
BIBLIOGRAPHY ........................................ 33
APPENDIX A. CHEMICAL STRUCTURE OF STEROIDS ........ 36
APPENDIX B. GRAPHS AND T A B L E S ..................... 37
LIST OP TABLES
TABLE PAGE
I. NaCl Solutions................................ 37
II. Cortin Sample of Unknown O r i g i n ............. 38
III. Wilson C o r t i n ................................ 39
IV. Cholic A c i d ........................ 40
V. Desoxycholic A c i d ............................ 4l
VI. Dehydrocholic Acid............................ 42
VTI. Dehydrodesoxycholic Aci d ...................... 43
VTII. Commercial Cortins ............................ 44
IX. James Wilson1 s Cortins........................ 45
X. James Wilson1s Cortin with Ringerfs ......... 46
XI. Desoxycholic Acid in Ringer's............... 47
XII. James Wilson's Cortin— Dilution Experiment . . 48
LIST OF FIGURES
FIGURE PAGE
1. Preliminary Experiment........................ 49
2. Desoxycholic Acid and N a C l ................... 49
3. Dehydrocholic Acid and NaCl .................... 50
4. Dehydrodesoxycholic Acid and NaCl . . . . . . . 50
5. Concentration Ringerfs Solution ........... 51
6. Desoxycholic Acid in Ringer’s ................. 51
CHAPTER I
THE PROBLEM
Statement of the problem. To investigate the effects
of steroid compounds, as represented by certain bile salts
and water soluble extracts of adrenal cortical tissue, on
the osmotic behavior of red blood cells.
Importance of the study. In view of the beneficial
effects which have been achieved with cortical extracts on
various human ills, it becomes important to know the ways
in which these compounds influence living tissue. Changes
in membrane permeability are one important effect they
are thought to exert. If effects could be found with
steroid concentrations at low physiological levels, the
actions of cortical extracts and bile salts on clinical
anemias might be postulated*— since it has long been known
that red blood cells have diminished resistance to hemoly­
sis in acholuric jaundice and other hemolytic anemias, and
an increased resistance in other types of anemias and
jaundice (Creed)(Dacie, Lond, and Vaughan).
Preview of organization of thesis. Topics will be
taken up in the following order: (1) a brief review of
the most important papers on the subject to indicate where
2
the problem stands today; (2) presentation of the methods
used in this investigation; the reasons for their use, and
the shortcomings and experimental error of such systems;
the results of the experiment; (3) discussion and theore­
tical considerations; evaluation of results and comments;
and (4) conclusions and summary.
CHAPTER IX
REVIEW OP THE LITERATURE
The state of the problem when this project was
begun. There are several papers relating directly to the
topic under discussion.
In 1939 & German investigator, Raushwalbe, reported
on tests he made with swine red blood cells and cortin1
( ,!cortidynn), in hypotonic saline and hypotonic glucose
solutions. His NaCl solutions ran from .4 - .Q%, with
cortin concentrations of 1:1000. He found that hemolysis
in the controls (i.e. without cortin) started at .7$ NaCl,
and that cortin prevented any hemolysis at this concentra­
tion. All of the effects he mentions were gross enough
to be visible to the naked eye, although he states that he
checked his results on a colorimeter. At .6$ NaCl he
noticed marked hemolysis in the controls, but very little
in the cortin test solutions.
Interestingly enough, he found that when hypotonic
glucose was used in place of NaCl, just the reverse hap­
pened— cortin promoted rather than inhibited hemolysis.
In his summary, this worker reverses himself, and
1 An extract of the adrenal cortex containing the
active steroids of the tissue.
4
says that cortin is hemolytic with salt and protective
with hypotonic glucose. It is to be assumed that that
statement made in the body of his paper, along with the
supporting data, represents the facts a3 he found them.
Dr. Chester Hyman of the University of Southern
California ran some similar experiments in 1947. These
were preliminary studies only, and are not intended to be
the basis for any conclusion on this problem. Using washed
rabbit blood cells and various concentrations of cortin
(1:2000 - 1:1000,000) in NaCl solutions which, without
cortin, gave approximately 50$ hemolysis, he found the
cortin inhibited hemolysis up to as much as 50$ of the con­
trol value in some instances, thus tending to confirm the
experiments of Raushwalbe. Hyman determined the relative
hemolysis from the color of the supernatant fluids as read
on a Klett photocolorimeter.
In 1938, Berliner and Schoenheimer studied the
hemolytic and anti-hemolytic properties of bile acids and
sterols, and attempted to relate these effects to their
chemical structure. (See Appendix I for brief review of
steroid and sterol structural formulae.) Their systems
differed from those described above in that the controls
were isoplethecontic, and hence were non-hemolytic. The
test solutions were made up of this isoplethecontic