INTRODUCTION:

The pituitary gland is responsible for the production and secretion of a variety of hormones that exert significant influence over other endocrine organs¹. Stress hormones are mainly regulated through this gland. Usually, the stress signs, known as ‘General adaptation syndrome’, comprises of alarm reaction, resistance and exhaustion, in which the ‘fight or flight’ alarm phase is triggered in the brain for the release of adrenocorticotropic hormone (ACTH) from the pituitary. Later this ACTH causes the adrenal glands to secrete adrenaline, cortisol, and other stress hormones². Among the cellular constituents of the anterior pituitary gland are corticotrophs, in addition to somatotrophs, gonadotrophs, lactotrophs, and thyrotrophs. The ultrastructural characteristics of corticotrophs are distinguished by a comparatively lower quantity of secretory granules relative to the other cell types of the adenohypophysis³. These granules are notable for their diminutive number, non-uniform size ranging from 80 to 250nm, electrodense appearance, and their predominant localization at the peripheral region of the cytoplasm. Furthermore, corticotrophs tend to be fewer in number and exhibit irregular sizing when compared to the other cellular populations within the adenohypophysis⁴.

A typical eukaryotic cell is characterized by cell membranes that create distinct extracellular and intracellular compartments. The transport of materials across this lipid bilayer membrane is facilitated by vesicles and tubules⁵. Eukaryotic cells have nucleus within inside of the cell and comprise of proper organelles with boundaries⁶. Additionally, the cell membrane plays several critical roles, including cell signaling, attachment to the cytoskeleton, adhesion to extracellular structures, and the conductivity of ions⁷. The nucleus, the largest organelle within the cell, is encased in a double membrane, effectively isolating it from the cytoplasm⁸. As the primary reservoir for DNA, RNA, proteins, and chromosomes, the size of the nucleus can vary significantly, with its principal role being the regulation of gene expression⁹. Mitochondria, which are also surrounded by double membranes, are essential for cellular survival, primarily functioning in energy production. This energy-generating process generates reactive oxygen species (ROS), which, when present in excessive amounts, can lead to mitochondrial damage¹⁰. The dysfunction of mitochondria may be associated with various detrimental effects, including metabolic disorders, inflammation, protein accumulation, and oxidative stress¹¹_. Inflammed necrotic cells usually leads to vascular responses, migration and activation of leucocytes, and systemic reaction¹². In general, the hypothalamo– pituitary–adrenal (HPA) axis along with the sympathoadrenal system will govern metabolic responses in our everyday life, along with overwhelming demands that exist under conditions of chronic stress¹³.

The endoplasmic reticulum (ER) plays a vital role as an organelle essential for maintaining cellular homeostasis. The phenomenon of ER stress may occur due to the accumulation of misfolded proteins, which subsequently activates the unfolded protein response (UPR)¹⁴. In response to stress, cells can trigger various mechanisms, such as the unfolded protein response (UPR), DNA damage response, mitochondrial stress signaling, and autophagy, along with localized tissue and broader systemic stress responses¹⁵. Certain dietary practices can place stress on the body at a cellular level, significantly affecting individual health and influencing the overall morbidity and mortality of populations¹⁶. Given the critical role of dietary habits in today's world, we aim to investigate how different diets impact the ultra-morphology of corticotrophs, crucial components in the stress response system.

MATERIALS AND METHODS:

Animal work:

In a controlled animal housing facility maintained at a temperature of 22 degrees Celsius, thirty-five eight-week-old Sprague-Dawley rats were acclimatized under conditions of a 12hour light-dark cycle. Following a two-week acclimatization period, the rats were stratified into five distinct groups (n=7). Each group was administered a specific diet as outlined by Azwan (2021), alongside ad libitum access to tap water. The dietary classifications included a control group receiving standard rat chow, and experimental groups receiving high-fat (35% fat), high-protein (52% protein), high-sugar (96% sugar), and high-starch (83% carbohydrates) diets. These diets were formulated using vegetable oil, whey protein, table sugar, and rice flour, respectively. After an eight-week feeding, the rats were euthanized in a carbon dioxide chamber, the pituitary glands harvested and processed for transmission electron microscopy analysis.

Transmission electron microscope:

The pituitary gland was fixed in a 4% glutaraldehyde solution overnight, followed by thorough washing across three changes of 0.1M sodium cacodylate buffer. Subsequently, samples underwent post-fixation with 1% osmium tetroxide and were then dehydrated through ascending grades of ethanol. After the dehydration process, the samples were embedded in resin overnight and subsequently placed into a beam capsule, allowing for polymerization over a 24hour period. Once prepared, the samples were subjected to ultrathin sectioning and placed onto copper grids, followed by staining with uranyl acetate. Additional contrast staining was performed using lead citrate prior to drying, preparing the samples for examination with the transmission electron microscope (JEOL JSM-7600F FESEM). Animal care and use were managed in accordance with the guidelines and recommendations set by the FOM IACUC, University of Malaya (Ref: 2019-21114/UNIKL/R/KAMJ).

RESULTS AND DISCUSSION:

The utilization of electron microscope to witness subcellular organelles and their changes is an essential instrument in achieving and assessing a visual understanding of cellular pathology and their prospect solution¹⁷. The nuclear envelope comprises a lipid bilayer, characterized by an inner layer enriched with proteins, while its outer surface is seamlessly integrated with the endoplasmic reticulum (ER). The primary function of the nuclear envelope is to physically separate and protect the deoxyribonucleic acid (DNA) from the cytoplasmic environment. Furthermore, molecular transport between the nucleus and cytosol is facilitated by the presence of nuclear pore complexes. Disruptions to the nuclear envelope, alongside mitochondrial swelling, membrane blebbing, nuclear fragmentation, and vacuole formation, constitute significant ultrastructural alterations associated with apoptosis¹⁸.

Figure 1. Corticotroph at 2000X magnification for 5 different diet groups. a – Control, b – high-fat, c – high-protein, d – high-sugar and e – high-starch.

No gap junctions were observed in all the exhibits observed in Figure 1. The nuclei of the corticotrophs exhibit an adequately homogeneous distribution of chromatin, except for the high-fat group (b), where a shrunken nucleus with peripheral aggregation of dense heterochromatin is evident. Numerous swollen mitochondria are present in both the high-fat (b) and high-sugar (d) groups, while the high-starch (e) group displays only a limited number of swollen mitochondria. In contrast, the control group (a) and the high-protein group (c) show minimal to no swollen mitochondria. Cytoplasmic protrusions are absent across all exhibits.

Figure 2. Nuclear membrane of corticotrophs at 10000X magnification.

In Figure 1, a notable reduction in the size of the nucleus in the high-fat group compared to the other groups is observed. However, various abnormalities pertaining to the nuclear envelope in both the high-fat and high-sugar groups are depicted in Figure 2, where the integrity of the smooth, double-layered nuclear membrane (indicated by red arrows) is clearly demonstrated in the control (a), high-protein (c), and high-starch (e) groups. Conversely, the high-fat (b) and high-sugar (d) groups exhibit a less distinct nuclear membrane. Furthermore, the high-fat group (b) displays pronounced clumping of condensed chromatin at the nuclear margins. Notably, the nuclear envelope in both groups appears indistinct, with the high-fat group exhibiting pronounced blebbing deformations. One study posits that nucleoplasmic pressure may exert force against the inner nuclear membrane, resulting in the formation of blebs, which are devoid of chromatin and consist solely of nucleoplasm ¹⁹. An alternative hypothesis suggests that deformation of the nucleus, accompanied by a reduction in volume, creates an osmotic pressure differential that leads to the development of blebs²⁰. These blebs are observed to form, rupture, and reform repeatedly, particularly during episodes of nuclear deformation, leading to interactions between nuclear and cytoplasmic materials, as the nuclear envelope intermittently opens and closes. Despite these recent observations, the precise mechanisms underlying their formation remain unresolved²¹.

Glycaemic level fluctuations, hypoxia, metabolic and oxidative stress conditions alter the shape and size of mitochondria and its cristae²². Mitochondrial swelling mechanism is illustrated by the inner mitochondrial membrane opening its non-selective permeability transition pores (PTP) whenever the mitochondria is threatened by adenosine triphosphate (ATP) depletion, oxidative stress, elevated calcium levels, increased reactive oxygen species (ROS) accumulation, and metabolic stress. Other factors such as high colloidal osmotic pressure within the mitochondrial matrix, ATP hydrolysis and depolarization of mitochondrial membrane contribute to mitochondrial swelling when chronic opening of PTP takes place²³.

Figure 3. Mitochondria of corticotrophs at 10000X magnification.

Mitochondrial swelling is prominently observed in groups subjected to high-fat, high-sugar, and high-starch diets, as illustrated in Figure 3. The mitochondria of the high-fat (b), high-sugar (d) and high-starch (e) groups are visibly swollen, and their cristae architecture disrupted and pushed just under the mitochondrial membrane. In the high-fat (b) and high-sugar (d) groups, the irregular shaped, highly damaged mitochondria are arranged together in clusters. These mitochondria appear to have a scattering of flocculant matrix and electron-dense deposits. The mitochondria of the control (a) and high-protein (c) group on the other hand appear oval or rounded in shape, not swollen and cristae structural integrity intact. The rupture of the outer mitochondrial membrane, resulting from this swelling, facilitates the release of pro-apoptotic proteins into the cytosol, consequently leading to either mitochondria-mediated necrosis or apoptosis²⁴. Pathological conditions associated with oxidative stress, including diabetes, hypoxia, and infarction-reperfusion, frequently manifest excessive mitochondrial swelling as a characteristic indicator²⁵. Numerous studies indicate that dietary components adversely affect mitochondrial function, primarily through mechanisms involving oxidative stress and reduced expression of antioxidant genes. Similarly, endoplasmic reticulum swelling was observed as shown in Figure 4, where swollen endoplasmic reticulum (red arrows) was observed in high-fat (a) and high-sugar (b) groups.

Figure 4. Showing swollen endoplasmic reticulum (red arrows) in corticotroph of high-fat (a) and high-sugar (b) groups at 10000X magnification. The control, high-protein and high-starch groups showed normal endoplasmic reticulum formation.

High-sugar diet has been linked to several detrimental effects, including protein glycosylation, accumulation of lipids, increased levels of reactive oxygen species (ROS), and alterations in membrane viscosity²⁶. Additionally, sustained elevated glucose levels have been associated with the upregulation of genes involved in mitophagy²⁷. Research investigating a low-protein diet indicated that the introduction of a high-fat ingredient resulted in swelling of multiple organelles, including mitochondria²⁸. Disruption of lipid and carbohydrate homoeostasis results in the development of chronic metabolic diseases like diabetes and atherosclerosis²⁹. Notably, within a mere two weeks of administration of a high-fat diet, a decline in brain electron transport chain proteins was detected, adversely affecting mitochondrial energy metabolism and morphology³⁰.

Additionally, mitochondrial swelling, along with diminished respiration levels in liver mitochondria, was reported in a high-fat diet study ³¹. Correlatively, mitochondrial disfigurement and dysfunction were linked to aberrant glucose metabolism induced by a high-fat diet³². A considerable body of animal studies has established the high-fat diet as a catalyst for metabolic disorders, demonstrating a detrimental impact on glucose tolerance, which subsequently leads to insulin resistance. The effect of stress related with triglycerides was due to discharge of corticosterone from adrenal cortex causing mobilization of lipids from adipose tissues³³. Moreover, it has been observed that a high-fat diet anomalously opens the permeability transition pore (PTP), resulting in mitochondrial inflammation through more reactive oxygen species (ROS) production, and disruption of mitochondrial physiology³⁴. The mechanism of oxidative stress usually through increased ROS formation caused by pro-oxidant enzymes like NADPH oxidase, mitochondrial oxidase, xanthine oxidase and nitric oxide synthase. In addition to ROS formation, decreased activity of antioxidant system also exist³⁵. Oxygen free radicals result in lipid peroxidation and causes damage to cell membranes with premature aging and cell death³⁶. Diet deficient in omega-3 fatty acids contribute to neuro degenerative inflammation³⁷. A study examining the effects of a high-sugar diet, which comprised 68% carbohydrates, revealed mitochondrial swelling and a disrupted cristae architecture within the gastrocnemius muscle cells of rodents³⁸. Research indicates that diabetic individuals experience complications and tissue damage resulting from a chronic hyperglycemic environment, causing oxidative stress³⁹. In a metabolic manuscript, it was proposed that excessive production of reactive oxygen species (ROS), activation of the hexosamine and polyol glycolytic pathways, and increased protein glycation are significant consequences of persistently elevated blood glucose levels⁴⁰. Within the central nervous system, increased redox reactions coupled with oxidative stress contribute to the progression of neurodegenerative diseases like Parkinson’s disease and Alzheimer’s disease, along with various cognitive impairments. These conditions have been extensively linked to high-sugar diets⁴¹. Notably, in the context of the brain, a high-sugar diet has been shown to adversely affect neurogenesis and neurotransmission activity⁴². In contrast, diet high in fruits and vegetables is associated with reduced effect of cancer and cardiovascular disease, along with age related disease⁴³. Among the various brain systems, the hypothalamus is particularly susceptible to the deleterious effects of a high-sugar diet when compared to the larger cerebral cortex⁴⁴. Consequently, our findings indicate that damage inflicted upon the corticotroph at the subcellular level, particularly due to high-fat and high-sugar dietary patterns, aligns with contemporary literature and evidence regarding oxidative stress and its detrimental effects.

CONCLUSION:

There exists a significant correlation between specific dietary patterns and alterations in the ultrastructure of corticotrophs. It is posited that diets high in fats and sugars exert considerable stress on the subcellular morphology and physiology of corticotrophs. It is imperative that further intensive genetic and molecular research be undertaken to elucidate and substantiate these findings.

CONFLICT OF INTERESTS:

The authors have no conflict of interest to declare.

ACKNOWLEDGEMENT:

This research is financed by the Malaysian Ministry of Higher Education (MOHE) via the Fundamental Research Grant Scheme (FRGS) with the grant listing number (FRGS/1/2018/SKK08/UNIKL/03/1).

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Received on 05.05.2025 Revised on 01.09.2025

Accepted on 06.11.2025 Published on 20.05.2026

Available online from May 25, 2026

Research J. Pharmacy and Technology. 2026;19(5):1955-1960.

DOI: 10.52711/0974-360X.2026.00280

This work is licensed under a Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International License. Creative Commons License.