The effect of Prednisolone on Live and Kidney of Newborn Rats

Dr. Lina M. Ali

Email: linaqahtan11@gmail.com
Phone: 07901297382
Abstract

This study aimed to examine the effects of prednisolone on the liver and
kidney of neonatal rats. Female rats were divided into four groups: G1 received
5 mg/kg of prednisolone, G2 received 15 mg/kg, G3 received 25 mg/kg, and G4
served as the control group and received a saline solution. The female rats were
mated, and embryos and fetuses were retrieved for analysis.
The study examined the effects of prednisolone on neonatal rats by
assessing their weight, length, and external morphological changes after
receiving prednisolone injections. The results revealed that the middle and high
dose-treated groups (G1, G2, and G3) experienced significant adverse effects on
the liver and kidney of neonatal rats.
These findings highlight the specific impact of prednisolone administration
during pregnancy on the liver and kidney of neonatal rats, raising concerns
about the potential risks associated with its use and its effects on neonatal
health.
This research contributes to a better understanding of the effects of
prednisolone on neonatal rats and emphasizes the importance of considering the
potential risks associated with its use during pregnancy.
‫خالصة‬
.‫هدفت هذه الدراسة إلى فحص تأثيرات بريدنيزولون على الكبد والكلى لدى الجرذان حديثي الوالدة‬
15 ‫تلقت‬G2 ، ‫ كجم من بريدنيزولون‬/ ‫ مجم‬5 ‫تلقت‬: G1 ‫تم تقسيم إناث الفئران إلى أربع مجموعات‬
‫ تم تزاوج‬.‫ خدمت كمجموعة تحكم وتلقت محلول ملحي‬G4 ‫ و‬، ‫ كجم‬/ ‫ مجم‬25 ‫تلقت‬G3 ، ‫ كجم‬/ ‫مجم‬
.‫ وتم استخراج األجنة واألجنة لتحليلها‬،‫إناث الفئران‬
‫فحصت الدراسة آثار بريدنيزولون على الفئران حديثي الوالدة من خالل تقييم وزنهم وطولهم‬
‫ أظهرت النتائج أن المجموعات المعالجة‬.‫والتغيرات المورفولوجية الخارجية بعد تلقي حقن بريدنيزولون‬
‫ تعرضت لتأثيرات سلبية كبيرة على الكبد والكلى لدى‬G3) ‫ و‬G2 ‫( و‬G1 ‫بجرعات متوسطة وعالية‬
.‫الفئران حديثي الوالدة‬
‫تسلط هذه النتائج الضوء على التأثير المحدد إلعطاء بريدنيزولون أثناء الحمل على كبد وكلى‬
‫ مما يثير مخاوف بشأن المخاطر المحتملة المرتبطة باستخدامه وتأثيراته على صحة‬،‫فئران حديثي الوالدة‬
.‫األطفال حديثي الوالدة‬
‫يساهم هذا البحث في فهم أفضل لتأثيرات بريدنيزولون على الفئران حديثي الوالدة ويؤكد على‬
.‫أهمية النظر في المخاطر المحتملة المرتبطة باستخدامه أثناء الحمل‬
Keywords: Glucocorticoids, Malformation, Newborn Rats, Prednisolone
Introduction

Pregnancy and fetal health have always presented researchers with

challenges, particularly in understanding the positive and negative
pharmacological effects (Christine et al., 2016).

Prednisolone, commonly prescribed during pregnancy to treat asthma,

autoimmune diseases, skin conditions, and inflammatory disorders such as
systemic lupus erythematosus (SLE/lupus) and rheumatoid arthritis (RA), has
shown practical anti-inflammatory effects and the ability to reduce swelling and
joint pain (Zimmerman et al., 2017).

Long-term and high-dose administration of Prednisolone has been

associated with significant effects on the liver, including acute liver injury that
can lead to acute liver failure and even death. However, it is important to note
that hepatic complications of Prednisolone usually arise from underlying liver
disease rather than direct drug hepatotoxicity (Chitturi et al., 2020).

Prednisolone therapy can induce hepatic steatosis and hepatic

enlargement, which may not be clinically apparent in adults. These effects can
occur rapidly and are typically reversible upon discontinuation. High doses and
long-term use have been linked to the exacerbation or expansion of
nonalcoholic steatohepatitis, characterized by elevated serum aminotransferase
levels and liver histology resembling alcoholic hepatitis with steatosis. Factors
such as alcoholism, malnutrition, pregnancy, or drug therapy can contribute to
this disturbance in liver cell metabolism (Schimmer et al., 2019).

Additionally, Prednisolone in high doses has been associated with hepatic

glycogenosis, which manifests as homogeneous liver cell appearance and strong
glycogen staining. Glycogenosis can be associated with hepatomegaly in
children and elevated serum aminotransferase levels, while alkaline phosphatase
levels may show minimal changes. Fortunately, glycogenosis is usually
asymptomatic and does not progress to chronic liver injury, cirrhosis, or acute
liver failure. Although commonly observed in patients with poorly controlled
type 1 diabetes, it can also occur acutely in patients initiated on high-dose
Prednisolone (Chitturi et al., 2020).

This research aimed to investigate the effect of prednisolone on liver

function and tissue in pregnant rats.

Materials and methods

The study utilized twenty female Sprague-Dawley rats weighing between

175 and 250 g, obtained from the Iraqi Center for Cancer Research / Al-
Mustansiriya University. The rats were housed in metal cages in the animal
house of the College of Science for Women / Baghdad University, under
controlled conditions of a temperature of 25°C. They were provided with ad
libitum access to water and food.

The female rats were randomly assigned to four groups, with five rats in
each group. Group G1 received an intraperitoneal injection of 5 mg/kg of
Prednisolone, Group G2 received 15 mg/kg, Group G3 received 25 mg/kg, and
Group G4 served as the control group and received distilled water.

To determine the stage of estrus, vaginal smears were collected from the
rats. Every two females were paired with one male in cages, and the presence of
a vaginal plug confirmed successful mating, indicating day 0 of pregnancy.
After mating, the male and female rats were separated to ensure accurate
gestational timing.
 Anesthesia was administered to the pregnant rats via intramuscular
injection of a mixture of Ketamine (90 mg/kg body weight) and Xylazine (10
mg/kg body weight). A midline incision was made in the abdomen to expose the
uterine horns. Hysterectomy was performed to retrieve the embryos and fetuses
from the placental sacs. The extra-embryonic membranes were removed, and
the embryos were rinsed with normal saline before being individually
examined.

The injected animals were sacrificed twelve hours after the final
administration of Prednisolone for the histological study of liver and kidney.
The results revealed that the middle and high dose-treated groups (G1, G2, and
G3) experienced significant adverse effects on the liver and kidney of neonatal
rats, compared to the control group.

This experimental design allowed for the assessment of the effects of

Prednisolone at different doses on the liver and kidney tissue of neonatal rats.
The study findings contribute to a deeper understanding of the impact of
Prednisolone administration during pregnancy.

Embryo obtaining

At 18 days of gestation, the pregnant rats were sacrificed, and the

embryos were carefully extracted from uterus of the pregnant rats for
histological examination, then some of the neonates were then transferred and
fixed in a solution of 10% formalin saline for 24-48 hours, followed by a
transfer and fixation in 70% ethanol solution.

This process involved assessing the liver and kidney sectioning by

microtome device for the neonatal rats in each group. Group G1 received an
intraperitoneal injection of 5 mg/kg of Prednisolone, Group G2 received 15
mg/kg, Group G3 received 25 mg/kg, while Group G4 served as the control
group and received distilled water.
 Sections of neonatal liver and kidney were examined under light
microscope to observe histological changes in all different doses (5, 15, and 25
mg/kg) of prednisolone compared to control group.

Result

The present histological study of liver and kidney tissue of the control
group (T1) showed normal hepatocytes arrange as plates around the central vein
with sinusoid between the liver plates (fig. 1, 2, 3, and 4)

Figure 1 Sections of neonatal liver of control group (H&E, 100x). A- Hepatocytes

surrounding a central vein C.V (black arrow). B- Hematopoietic components (black arrow),
hepatic sinusoidal (S), and megakaryocytes (red arrow). C- immature bile ductules (black
arrow) and portal vein (PV).
Figure 2: Sections of neonatal liver exposed maternally to Prednisolone in (G1) (H&E, 400x).
A- Low dose group (5 mg/kg) shows sinusoidal congestion (black arrow) while other filled
with mild haemopoietic cells (red arrow). B- vacuolar degeneration (arrow) with absence of
haemopoietic component. C- massive dilation of C.V. with slight myeloid cells infiltration in
the sinusoids (arrow).

Figure 3: Sections of neonatal liver exposed maternally to Prednisolone in (G2) (H&E). A-

middle (15 mg/kg) dose group shows empty spaces (arrow) in the hepatic parenchyma 100x.
B- fibroplasia (red arrow) with lymphocytes infiltration (black arrow). 400x. C- necrotic foci
(asterisk) with sinusoidal congestion and dilation (black arrow). 400x.
Figure 4: Sections of neonatal liver exposed maternally to Prednisolone in (G3) (H&E). A-
high dose group (25 mg/kg) shows wide hemorrhagic area including subscapular (black
arrow) 100x. B- necrosis with prominent of haemopoietic cells (black arrow) 400x. C-
individualization of hepatocytes with more acidophilic appearance (black arrow) 400x.

Figure 5: Sections of neonatal kidney of control group (H&E). A- subcapsular nephrogenic

zone (N), comma-shaped body (white arrow), S-shaped body (black arrow), and glomerulus
 (red arrow). 400x. B- medullar portion (M), ureteric bud (U), and immature glomerulus
(arrow) .400x. C- renal cortex (C) and medullar portion (M). 100x.

Figure 6: Sections of neonatal kidney exposed maternally to Prednisolone in (G1) (H&E,

400x). A- Low dose group (5 mg/kg) shows severe degeneration of some renal vesicles
(arrow) in subcapsular nephrogenic zone (N). B- tubular dilation and glomerular tuft
shrinkage. C- focal tubular necrosis (asterisk).

Figure 7: Sections of neonatal kidney exposed maternally to Prednisolone (G2) (H&E,

400x). A- middle dose group (15 mg/kg) shows blood vessel congestion and dilation (red
arrow) with intratubular edema (black arrow). B- slight distention of Bowman’s space with
edema (black arrow), rupture in dilated tubules (red arrow). C- subscapular hemorrhage (red
arrow) and disruption of cortical tubules (black arrow).
 Figure 8: Sections of neonatal kidney exposed maternally to Prednisolone (G3) (H&E). A-
high dose group (25 mg/kg) shows glomerulus with massive hypercellularity in deeper part
of renal cortex (arrow). 400x. B- cortical cyst formation (asterisk). 100x. C- dilation of
Bowman’s space with edematous substance crescent formation (arrow) and severe loss of
surrounding parenchyma (asterisk). 400x.

Discussion

The fetal liver is a target organ that responds to maternal exposure to different
environmental agents, including prednisolone. Prednisolone readily enters the
liver through the mononuclear phagocytic system. In a study involving pregnant
rats exposed to varying doses of prednisolone during the G1, minor time-
dependent histological changes were observed in the neonatal liver compared to
other treatment and control groups. These changes may be associated with the
formation of the visceral yolk sac and the early stages of placenta formation,
allowing prednisolone transfer to the developing embryo.

In G2, more pronounced histological changes, such as parenchymal loss

and focal necrosis, were observed in the hepatic tissue of neonates. These
changes can be attributed to the prolonged accumulation of prednisolone in
female rats prior to mating and during early pregnancy. Previous studies have
shown that prednisolone residues persist in the bodies of mature rats for an
extended period and decrease over time in recipient organs. Additionally,
prednisolone has the ability to accumulate in fetuses by crossing the placental
barrier.

In G3, the histological alterations in the neonatal liver were more significant
with a middle dose of prednisolone compared to other groups. This can be
explained by the tendency of prednisolone to agglomerate at high
concentrations, which reduces its transfer across the placental barrier and
decreases silver deposition in the fetal liver. The metabolic function of
hepatocytes during this stage leads to increased oxidative stress, suppressed
antioxidant defense, lipid peroxidation, and cell apoptosis.

Histological changes in the neonatal liver in G3 were also observed, primarily

due to the accumulation of prednisolone in the late stages of pregnancy. The
decreased thickness of the placental barrier and increased fetal capillaries
facilitate the transport of prednisolone, resulting in its accumulation in the fetus.
This accumulation leads to hepatic lesions, including hepatocytes with fatty
droplets, atrophy, necrosis, apoptosis, and disruption of hepatic cords.

The effects of prednisolone on hemopoietic cells were also observed. The

fetal liver, which is the main organ for hemopoietic cell production during the
fetal development stage and the first week of neonatal rats, showed a depletion
or absence of hemopoietic elements, lymphocytolysis, and decreased erythroid
and myeloid cells. This may be due to the impact of prednisolone on the
proliferation and differentiation of hemocytoblasts and final hemopoietic
products, leading to necrosis or apoptosis.

Studies on human primary erythroid cells exposed to prednisolone showed

increased hemolysis and apoptosis, indicating elevated oxidative stress and ROS
levels. Prednisolone were also found to inhibit erythropoiesis in the fetal liver
by interfering with RNA polymerase and repressing RNA transcription,
resulting in reduced cell proliferation.

Overall, the exposure of pregnant rats to prednisolone led to histological

changes in the fetal liver, affecting various aspects such as tissue morphology,
hemopoietic cells, and metabolic function.

On the other hand, our study focused on the effects of prednisolone on the
developing kidneys of newborn rats when exposed to varying doses. The
histological changes in the neonatal kidneys were observed to be time-
dependent, with minor alterations during the first week and more pronounced
changes during the second and third weeks. The transfer of prednisolone to the
embryo was facilitated by the visceral yolk sac and placenta formation. The
developing kidneys showed higher susceptibility to prednisolone during the
third week, corresponding to the full development and functional maturation of
nephrons. TEM analysis revealed the distribution of prednisolone across the
placenta and their influence on fetal toxicity. Histopathological changes
included the accumulation of the renal structures, leading to cytoplasmic and
nuclear damage, disruption of cellular organelles, and induction of necrosis and
apoptosis. The alterations observed in the neonatal kidneys during the second
week included glomerular and tubular necrosis, loss of renal parenchyma, and
subcapsular hemorrhage. The dysregulation of cell proliferation and apoptosis,
as well as changes in cell polarity and interaction, were associated with cortical
cyst formation. The study highlights the potential nephrotoxic effects of
prednisolone on the developing kidneys and the importance of considering the
stage of feto-placental development during nanoparticle exposure.

Conclusion

In conclusion, the histopathological examination of the liver in newborn

rats exposed to Prednisolone revealed significant alterations. This included
disarrangement of hepatocytes, congestion in central veins, sinusoid expansion,
necrosis, and glycogen deposition. These histological changes indicate the
potential adverse effects of Prednisolone on the liver of neonatal rats.

The findings suggest that Prednisolone administration during pregnancy

may disrupt normal liver morphology and function in newborn rats. These
histopathological alterations raise concerns about the potential impact on fetal
development and overall liver health.

Further research is necessary to elucidate the underlying mechanisms

responsible for these histopathological changes and to determine their long-term
implications. Understanding the specific effects of Prednisolone on the liver of
neonatal rats will contribute to a better understanding of the risks associated
with its use during pregnancy.

Considering the observed histopathological findings, caution should be

exercised when prescribing Prednisolone to pregnant individuals. Healthcare
professionals should carefully weigh the potential risks and benefits before
initiating treatment with Prednisolone during pregnancy. Additionally,
strategies should be explored to mitigate the potential adverse effects on liver
health in newborns exposed to Prednisolone.

Overall, the histopathological results of this study highlight the need for
further investigation and monitoring of liver health in offspring exposed to
Prednisolone, aiming to ensure the optimal management and well-being of
neonates.

References: -

1. Vogt, M.; Derendore, H.; Kramer, J.; Junginger, H.E.; Midha, K.K.; Shah,
V.P.; Stavchansky, S.; Deessman, J.B., and Barends, D.M. (2007).
Biowaiver Monographs for Immediate Release Solid Oral Dosage Forms:
Prednisolone. J. of Pharmaceutics. Sci. , vol. 96 , no. 1, Jan, 27 – 37.
2. Ramsahoye, B.H.; Davies, S.V., and Sandeman, D. (1995). Prednisolone.
International J. of Pharmaceutics., vol.120 , n: 30 , June, 197 – 204.

3. Francisco, G.E.; Honigberg, I.L., and Stewart, J.T. (1984). In vitro and in
vivo bioequivalence of commercial prednisone tablets. Biopharm. Drug

Dispos. 5 : 335 – 344.

4. Kaiser, H., and Kley, H.K. (2002). Cortisontherapie. Corticoid in Klinik und
Praxis.

11. Auflage. Georg Thieme Verlag, Stuttgart, Germany.

5. Richard, D.H. and Mary, Y.J.M. (2006). Lippicotts

Illustrated Review: Pharmacology. Ed., Lippicott Williams and
Wilkins. Awolters Kluwer Company.

6. Pickup, M.E. (1979). Clinical pharmacokinetics of prednisone and

prednisolone. Clin. Pharmacokin. 4 : 111 – 128.

7. Petereit, L.B., and Meikle, A.M. (1977). Effectiveness of prednisolone

during phenytoin therapy. Clin. Pharmacol. Ther. 22 : 912 – 916.

8. Fekety, R.; Gorbach, S.L.; Bartlett, J.G., and Blacklow, N.R. (1992).
Infections associated with corticosteroids and immunosuppressive therapy
in : Infectious Diseases. Philadelphia: WB Saunders Company.1050 - 1.

9. Stuck, A.E.; Minder, C.E., and Frey, F.J. (1989). Risk of infectious
complications in patients taking glucocorticoids. Rev. Infect. Dis., 11(6)
:954 - 63.

10. Rose, J.Q.; Yurchak, A.M., and Jusko, W.J. (1981). Dose dependent
pharmacokinetics of prednisone and prednisolone in man. J. Pharmacokinet.
Biopharm. 9 : 389 – 417.
11. Pickup, M.E.; Lose, J.R.; Leatham, P.A.; Rhind, V.M.; Wright, V., and
Downie, W.W. (1977). Dose dependent pharmacokinetics of prednisolone.
Eur. J. Clin. Pharmacol. 12 : 213 – 219.

12. Nugent, C.A.; Eik-nes, K., and Tyler, F.H. (1959). A comparative study of
the metabolism of hydrocortisone and prednisolone. J. Clin. Endocrinol.
Metab. 19 : 526 – 534.

13. Hartiala, J. (1976). Steroid metabolism in adult lung. Clin. Pharmacol. Ther.
25 : 571 – 578.

14. Frey, B.M., and Frey, F.J. (1990). Clinical pharmacokinetics of prednisone
and prednisolone. Clin Pharmacokinet 19:126–146.

15. Ramsahoye, B. H.; Davies, S.V.; Gaylani, N.F.L.; Sandeman, D., and
Scanlon, M.F. (1995). The mineralocorticoid effects of high dose
hydrocortisone. B.M.J. , vol. 310 : 656 – 657.

16. Troels, W.; Ole, H.; Thorbjorn, G., and Hendrik, V. (2000) . Effects of
Budesonide and Prednisolone on hepatic kinetics for urea synthesis. Journal
of Hepatology. , vol. 33, n 4, pp. 549 – 554.

17. Grofte, T.; Jensen, D.S.; Gronbaek, H.; Wolthers, T.; Jensen, S.A.;Tygstrup,
N., and Vilstrup, H.(1998). Effect of growth hormone on steroid-induced
increase in ability of urea synthesis and urea enzyme mRNA levels. Am. J.
Physiol. Endocrinol. Metab. , vol. 275 , Issue 1, E79 – E86 , July.

18. Ito, S.; Ohkoshi, S.; Aoyagi, T.; Suzuki, K.; Takahashi, T.; Nomoto, M.;
Nakano, M.; Arakawa, M.; Asakura, H., and Gejyo, F. (2003). A Patient
with Takayasu 's Arthritis Treated with Corticosteroids who developed
primary biliary cirrhosis. Internal Medicine. , vol. 42 : 443 - 445.

19. Scheuer, p. (1967). Primary biliary cirrhosis. Proc. R. Soc. Med. 60 : 1257 –
1260.
20. Reibmans, and Frankel, S. (1957). Am. J. Clin. Path., 28 : 56

21. Kind, P.R.N., and King, E.J. (1954). Estimation of plasma phosphatase by
determination of hydrolyzed phenol with amino – antipyrine. J.Clin. Path., 7
: 322 -326.

22. Wills, M.R., and Savory, J. (1981). Biochemistry of renal failure. Am.Clin.
Lab. Sci., vol.11, n.4p, 292 – 299.

23. Al–Mohammed, K.M.; Al–Rawi,Y.M.A., and Al– Morani,W.K. (1986).

Principles of Statistics. Al – Mousl University.

24. Kelly, L.(2004). Essential of Human Physiology for Pharmacy. CRC Press
Pharmacy Education Series. Baco Raton London, New York, Washington,
D.C.

25. Pocock, G., and Richards, C.D. (2004). Human Physiology, The Basis of
Medicine. 2nd ed. Oxford Medical Publications. Oxford University Press ,
Inc., New York.