Single Cell Layer Research Articles

Characterizing the role of pulmonary veins in pulmonary hypertension

Pulmonary veins (PVs) transport oxygenated blood from lungs back to the heart. Small PVs can play a crucial role in the pathogenesis of pulmonary hypertension. Studies have also shown PV remodeling in pulmonary hypertension and heart failure. However, unlike pulmonary arteries (PAs), which have been a major focus of pulmonary vascular studies, the signaling mechanisms in PVs have not been investigated under normal conditions and in pulmonary hypertension. We hypothesized that the vascular wall composition and cellular signaling mechanisms in PVs are distinct from PAs. We imaged 4th order PVs (~ 50 mm) from C57BL6 mice using spinning disk confocal imaging along the z-axis. In direct contrast to PAs, where a single endothelial cell (EC) layer is surrounded by alpha actin-positive smooth muscle cells (SMCs), PV walls did not show SMCs. Cardiac troponin C staining of PVs identified cardiac myocyte-like (CML) cells with striations surrounding the EC layer. The CML layer was not observed in small PAs, vena cava, and mesenteric veins. Confocal calcium imaging showed spontaneous calcium signals in CML cells from PVs that were inhibited by ryanodine (1 mM), a ryanodine receptor inhibitor. Cannulated PVs in pressure myography experiments showed spontaneous pulsatile constrictions, which were also inhibited by ryanodine. Collectively, our results suggest that ryanodine receptor calcium signals in CML cells underlie spontaneous contractions in small PVs. Our findings also indicate that signaling mechanisms in CML cells could be potential targets for therapy in pulmonary hypertension. This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

The surface ectoderm exhibits spatially heterogenous tension that correlates with YAP localisation during spinal neural tube closure in mouse embryos

The single cell layer of surface ectoderm (SE) which overlies the closing neural tube (NT) plays a crucial biomechanical role during mammalian NT closure (NTC), challenging previous assumptions that it is only passive to the force-generating neuroepithelium (NE). Failure of NTC leads to congenital malformations known as NT defects (NTDs), including spina bifida (SB) and anencephaly in the spine and brain respectively. In several mouse NTD models, SB is caused by misexpression of SE-specific genes and is associated with disrupted SE mechanics, including loss of rostrocaudal cell elongation believed to be important for successful closure. In this study, we asked how SE mechanics affect NT morphology, and whether the characteristic rostrocaudal cell elongation at the progressing closure site is a response to tension anisotropy in the SE. We show that blocking SE-specific E-cadherin in ex utero mouse embryo culture influences NT morphology, as well as the F-actin cable. Cell border ablation shows that cell shape is not due to tension anisotropy, but that there are regional differences in SE tension. We also find that YAP nuclear translocation reflects regional tension heterogeneity, and that its expression is sensitive to pharmacological reduction of tension. In conclusion, our results confirm that the SE is a biomechanically important tissue for spinal NT morphogenesis and suggest a possible role of spatial regulation of cellular tension which could regulate downstream gene expression via mechanically-sensitive YAP activity.

Imaging Recommendations for Diagnosis, Staging, and Management of Peritoneal Malignancies

AbstractPeritoneum is a serosal membrane lining the solid viscera and the hollow viscus of the abdomen and is made of a single layer of mesothelial cells. The most common primaries that spread to the peritoneum include gastrointestinal, ovarian, colorectal, and peritoneal metastases can be seen at some point during the disease course in 15 to 43%, 60 to 70% and 15 to 20% of patients, respectively. Other malignancies involving the peritoneum such as primary peritoneal carcinoma, peritoneal mesothelioma, peritoneal lymphomatosis, pseudomyxoma peritonei from low-grade appendiceal mucinous neoplasm, are far less common. The review strives to provide a framework for diagnosis and management of the disease.

Histological changes of the skin during postembryonic development of the crested newt Triturus ivanbureschi (Urodela, Salamandridae)

BackgroundAmphibian skin has been studied for many decades, especially the metamorphic changes in the skin of frogs. Less attention has been paid to salamander skin. Here, we describe changes in the skin structure during postembryonic development in a salamandrid species, the Balkan crested newt Triturus ivanbureschi. MethodUsing traditional histological techniques we examined the skin in the trunk region of three premetamorphic larval stages (hatchling, mid larval and late larval) and two postmetamorphic stages (juvenile, just after metamorphosis, and adult). ResultsIn larval stages, skin consists only of the epidermis, which gradually develops from the single epithelial cell layer in hatchlings, to a stratified epidermis with gland nests and characteristic Leydig cells at the late larval stage. During metamorphosis, Leydig cells disappear, and the dermal layer develops. In postmetamorphic stages, skin is differentiated on stratified epidermis and the dermis with well-developed glands. Three types of glands were observed in the skin of the postmetamorphic stages: mucous, granular and mixed. Gland composition appears to be stage- and sex-specific, with juveniles and adult female being more similar to each other. In juveniles and adult female, there are a similar proportion of glands in both dorsal and ventral skin, whereas in adult male granular glands dominated the dorsal skin, while mixed glands dominated the ventral skin. ConclusionOur results provide a baseline for future comparative research of skin anatomy in salamanders.

The Multifarious Effects of Various Glaucoma Pharmacotherapy on Corneal Endothelium: A Narrative Review.

Corneal endothelium is a single cell layer that is mainly responsible for maintaining corneal clarity. Endothelial damage secondary to toxicity, stress, or genetic predisposition are common and in conjunction with the low regenerative ability of the cells, making their preservation critical for maintaining visual acuity. Patients with glaucoma, who are estimated to be close to 80 million worldwide, have a plethora of reasons for developing endothelial damage, being exposed to a spectrum that extends from various medical and surgical interventions to the disease itself. The wide spectrum of glaucoma pharmacotherapy that has been recently extended by addition of newer classes of medications has been the focus of extensive research on its effects on corneal endothelium. Both basic and clinical research have attempted to shine a light on the complex mechanisms associated with the effects of glaucoma medication on corneal endothelium and to answer the important question as to whether these findings are clinically significant. The aim of this review is to summarize and present current literature of the various findings, both from invivo and invitro studies that have focused on the complex relationship between different classes of glaucoma medication and their effect on corneal endothelium.

Thymic cavernous haemangioma in a dog

A 12-year-old male neutered Australian Shepherd Dog was presented to Charlotte Animal Referral & Emergency with a history of a thoracic mass. On physical examination, physiological parameters were within the normal ranges, and a complete haemogram and serum biochemistry profile were unremarkable except for mild thrombocytopenia. A computed tomography scan revealed a 21 × 15 × 12.7 cm thoracic mass encompassing the cranial mediastinum and extending to the right caudal thorax. The mass was surgically removed and histopathological evaluation revealed that it comprised remnants of the thymus and a neoplasm composed of large blood-filled vascular spaces lined by a single layer of endothelial cells with mild anisocytosis and anisokaryosis. The neoplastic cells had diffuse strong immunolabeling for endothelial cell marker CD31. Multifocally, there were large cystic degenerated areas of thymic tissue lined by plump cytokeratin AE1/AE3-positive epithelial cells. Based on these findings, a diagnosis of thymic cavernous haemangioma (CH) was made. Thymic CH is rare in animals, with the only reported case in a cross bred cow. To our knowledge, this is the first case of a thymic CH in a dog.

Cross-linking porcine peritoneum by oxidized konjac glucomannan: a novel method to improve the properties of cardiovascular substitute material

The use of natural polysaccharide crosslinkers for decellularized matrices is an effective approach to prepare cardiovascular substitute materials. In this research, NaIO4 was applied to oxidize konjac glucomannan to prepare the polysaccharide crosslinker oxidized konjac glucomannan (OKGM). The as-prepared crosslinker was then used to stabilize collagen-rich decellularized porcine peritoneum (DPP) to construct a cardiovascular substitute material (OKGM-fixed DPP). The results demonstrated that compared with GA-fixed DPP and GNP-fixed DPP, 3.75% OKGM [1:1.5 (KGM: NaIO4)]-fixed DPP demonstrated suitable mechanical properties, as well as good hemocompatibility, excellent anti-calcification capability, and anti-enzymolysis in vitro. Furthermore, 3.75% OKGM [1:1.5 (KGM: NaIO4)]-fixed DPP was suitable for vascular endothelial cell adhesion and rapid proliferation, and a single layer of endothelial cells was formed on the fifth day of culture. The in vivo experimental results also showed excellent histocompatibility. The current results demonstrted that OKGM was a novel polysaccharide cross-linking reagent for crosslinking natural tissues featured with rich collagen content, and 3.75% OKGM [1:1.5 (KGM: NaIO4)]-fixed DPP was a potential cardiovascular substitute material.Graphical

Characterizing Epithelial Wound Healing In Vivo Using the Cnidarian Model Organism Clytia hemisphaerica.

All animal organs, from the skin to eyes to intestines, are covered with sheets of epithelial cells that allow them to maintain homeostasis while protecting them from infection. Therefore, it is not surprising that the ability to repair epithelial wounds is critical to all metazoans. Epithelial wound healing in vertebrates involves overlapping processes, including inflammatory responses, vascularization, and re-epithelialization. Regulation of these processes involves complex interactions between epithelial cells, neighboring cells, and the extracellular matrix (ECM); the ECM contains structural proteins, regulatory proteins, and active small molecules. This complexity, together with the fact that most animals have opaque tissues and inaccessible ECMs, makes wound healing difficult to study in live animals. Much work on epithelial wound healing is therefore performed in tissue culture systems, with a single epithelial cell-type plated as a monolayer on an artificial matrix. Clytia hemisphaerica (Clytia) provides a unique and exciting complement to these studies, allowing epithelial wound healing to be studied in an intact animal with an authentic ECM. The ectodermal epithelium of Clytia is a single layer of large squamous epithelial cells, allowing high-resolution imaging using differential interfering contrast (DIC) microscopy in living animals. The absence of migratory fibroblasts, vasculature, or inflammatory responses makes it possible to dissect the critical events in re-epithelialization in vivo. The healing of various types of wounds can be analyzed, including single-cell microwounds, small and large epithelial wounds, and wounds that damage the basement membrane. Lamellipodia formation, purse string contraction, cell stretching, and collective cell migration can all be observed in this system. Furthermore, pharmacological agents can be introduced via the ECM to modify cell:ECM interactions and cellular processes in vivo. This work shows methods for creating wounds in live Clytia, capturing movies of healing, and probing healing mechanisms by microinjecting reagents into the ECM.

Hesperidin enhances intestinal barrier function in Caco-2 cell monolayers via AMPK-mediated tight junction-related proteins.

The intestinal epithelium is a single-cell layer on the mucosal surface that absorbs food-derived nutrients and functions as a barrier that protects mucosal integrity. Hesperidin (hesperetin-7-rhamnoglucoside) is a flavanone glycoside composed of the flavanone hesperetin and the disaccharide rutinose, which has various physiological benefits, including antioxidative, anti-inflammatory, and antiallergic effects. Here, we used human intestinal Caco-2 cell monolayers to examine the effect of hesperidin on intestinal barrier function. Hesperidin-treated Caco-2 cell monolayers displayed enhanced intestinal barrier integrity, as indicated by an increase in transepithelial electrical resistance (TEER) and a decreased apparent permeability (Papp ) for fluorescein. Hesperidin elevated the mRNA and protein levels of occludin, MarvelD3, JAM-1, claudin-1, and claudin-4, which are encoded by tight junction (TJ)-related genes. Moreover, hesperidin significantly increased the phosphorylation of AMP-activated protein kinase (AMPK), indicating improved intestinal barrier function. Thus, our results suggest that hesperidin enhances intestinal barrier function by increasing the expression of TJ-related occludin, MarvelD3, JAM-1, and claudin-1 via AMPK activation in human intestinal Caco-2 cells.

The fungicide azoxystrobin causes histopathological and cytotoxic changes in the midgut of the honey bee Apis mellifera (Hymenoptera: Apidae).

Apis mellifera is an important bee pollinating native and crop plants but its recent population decline has been linked to the use of pesticides, including fungicides that have been commonly classified as safe for bees. However, many pesticides, in addition to direct mortality cause sublethal effects, including damage to target selective honey bee organs. The midgut is the organ responsible for the digestion and absorption of nutrients and the detoxification of ingested substances, such as pesticides. This study evaluated the histopathological and cytotoxic changes in the midgut of A. mellifera workers caused by the pesticide azoxystrobin. The limit-test was performed, and a 100 µg a.i./bee dose was administered orally and midgut analyzed with light and transmission electron microscopies after 24 h and 48 h of pesticide exposure. The midgut of the control bees has a single layer of digestive cells, with spherical nuclei, nests of regenerative cells, and the lumen coated with the peritrophic matrix. The bees fed on azoxystrobin showed morphological changes, including intense cytoplasm vacuolization and cell fragments released into the gut lumen. The protein detection test showed greater staining intensity in the nests of regenerative cells after 24 h of exposure to azoxystrobin. The occurrence of damage to the midgut in A. mellifera exposed to azoxystrobin indicates that although this fungicide has been classified as low toxicity for bees, it has sublethal effects in the midgut, and effects in other organs should be investigated.

Rab11 negatively regulates wingless preventing JNK-mediated apoptosis in Drosophila epithelium during embryonic dorsal closure.

Rab11, a small Ras like GTPase marking the recycling endosomes, plays instrumental roles in Drosophila embryonic epithelial morphogenesis where an array of reports testify its importance in the maintenance of cyto-architectural as well as functional attributes of the concerned cells. Proper Rab11 functions ensure a precise regulation of developmentally active cell signaling pathways which in turn promote the expression of morphogens and other physico-chemical cues which finally forge an embryo out of a single layer of cells. Earlier reports have established that Rab11 functions are vital for fly embryonic development where amorphic mutants such as EP3017 homozygotes show a fair degree of epithelial defects along with incomplete dorsal closure. Here, we present a detailed account of the effects of Rab11 loss of function in the dorso-lateral epithelium which resulted in severe dorsal closure defects along with an elevated JNK-Dpp expression. We further observed that the dorso-lateral epithelial cells undergo epithelial to mesenchymal transition as well as apoptosis in Rab11 mutants with elevated expression levels of MMP1 and Caspase-3, where Caspase-3 contributes to the Rab11 knockout phenotype contrary to the knockdown mutants or hypomorphs. Interestingly, the elevated expressions of the core JNK-Dpp signaling could be rescued with a simultaneous knockdown of wingless in the Rab11 knockout mutants suggesting a genetic interaction of Rab11 with the Wingless pathway during dorsal closure, an ideal model of epithelial wound healing.

Morphogenetic events influencing corneal maturation, development, and transparency: Light and electron microscopic study.

The development of the cornea is a fascinating process. Its dual origin involves the differentiation of surface ectoderm cells and the migration of mesenchymal cells of neural crest origin. This research aimed to demonstrate the morphogenesis of the rabbit cornea from fetal to postnatal life using light- and electron microscopy, and immunohistochemical analysis. There were 27 rabbit embryos and nine rabbits used. The rabbit cornea begins its prenatal development on the twelfth day of gestation. The surface ectoderm differentiates into the corneal epithelium on day 13. Intriguingly, telocytes were visible within the epithelium. The secondary stroma develops on the sixteenth day of gestation by differentiation of keratocytes. At the age of 2 weeks, the lamellae of collagenous fibers become highly organized, and the stroma becomes avascular, indicating that the cornea has become transparent. Bowman's membrane appears on day 23 of pregnancy and disappears on day 30. The Descemet's membrane appears at this time and continues to thicken postnatally. The corneal endothelium appears on the twentieth gestational day as double layer of flattened cells and becomes a single layer of cuboidal cells on day 30. The spaces between the endothelial cells resemble craters. VEGF immunohistochemical expression increases over the course of development, reaching its peak in the first week after birth before decreasing in all corneal layers and becoming negative in the stroma. In conclusion, numerous morphogenetic events contribute to corneal maturation and transparency, allowing the cornea to perform its vital functions.

Papillomatous breast lesions with atypical columnar cell features

AimsColumnar cell lesions (CCLs) are recognised breast cancer precursor lesions. Intraductal papillomas are usually lined by benign (polyclonal) cells. Although papillomas with monoclonal lesions (atypical ductal hyperplasia (ADH)/ductal carcinoma in...

Confocal Laser Scanning Imaging of Cell Junctions in Human Colon Cancer Cells.

The intestinal epithelium is formed by a single layer of cells. These cells originate from self-renewal stem cells that give rise to various lineages of cells: Paneth, transit-amplifying, and fully differentiated cells (as enteroendocrine, goblet cells, and enterocytes). Enterocytes, also known as absorptive epithelial cells, are the most abundant cell type in the gut. Enterocytes have the potential to polarize as well as form tight junctions with neighbor cells which altogether serve to ensure both the absorption of "good" substances into the body and the blockage of "bad" substances, among other functions. Culture cell models such as the Caco-2 cell line have been proved to be valuable tools to study the fascinating functions of the intestine. In this chapter we outline some experimental procedures to grow, differentiate, and stain intestinal Caco-2 cells, as well as image them using two modes of confocal laser scanning microscopy.

Strong spin-lattice entanglement in cobaltites

Strongly correlated electronic system contains strong coupling among multi-order parameters and is easy to efficiently tune by external field. Cobaltite (LaCoO<sub>3</sub>) is a typical multiferroic (ferroelastic and ferromagnetic) material, which has been extensively investigated over decades. Conventional research on cobaltites has focused on the ferroelastic phase transition and structure modulation under stress. Recently, researchers have discovered that cobaltite thin films undergo a paramagnetic-to-ferromagnetic phase transition under tensile strain, however, its origin has been controversial over decades. Some experimental evidence shows that stress leads the valence state of cobalt ions to decrease, and thus producing spin state transition. Other researchers believe that the stress-induced nano-domain structure will present a long-range ordered arrangement of high spin states, which is the main reason for producing the ferromagnetism of cobalt oxide films. In this paper, we review a series of recent researches of the strong correlation between spin and lattice degrees of freedom in cobalt oxide thin films and heterojunctions. The reversible spin state transition in cobalt oxide film is induced by structural factors such as thin-film thickness, lattice mismatch stress, crystal symmetry, surface morphology, interfacial oxygen ion coordination, and oxygen octahedral tilting while the valence state of cobalt ions is kept unchanged, and thus forming highly adjustable macroscopic magnetism. Furthermore, the atomic-level precision controllable film growth technology is utilized to construct single cell layer cobaltite superlattices, thereby achieving ultra-thin two-dimensional magnetic oxide materials through efficient structure regulation. These advances not only clarified the strong coupling between lattice and spin order parameters in the strongly correlated electronic system, but also provided excellent candidate for the realization of ultra-thin room temperature ferromagnets that are required by oxide spintronic devices.

A kidney proximal tubule model to evaluate effects of basement membrane stiffening on renal tubular epithelial cells.

The kidney tubule consists of a single layer of epithelial cells supported by the tubular basement membrane (TBM), a thin layer of specialized extracellular matrix (ECM). The mechanical properties of the ECM are important for regulating a wide range of cell functions including proliferation, differentiation and cell survival. Increased ECM stiffness plays a role in promoting multiple pathological conditions including cancer, fibrosis and heart disease. How changes in TBM mechanics regulate tubular epithelial cell behavior is not fully understood. Here we introduce a cell culture system that utilizes in vivo-derived TBM to investigate cell-matrix interactions in kidney proximal tubule cells. Basement membrane mechanics was controlled using genipin, a biocompatibility crosslinker. Genipin modification resulted in a dose-dependent increase in matrix stiffness. Crosslinking had a marginal but statistically significant impact on the diffusive molecular transport properties of the TBM, likely due to a reduction in pore size. Both native and genipin-modified TBM substrates supported tubular epithelial cell growth. Cells were able to attach and proliferate to form confluent monolayers. Tubular epithelial cells polarized and assembled organized cell-cell junctions. Genipin modification had minimal impact on cell viability and proliferation. Genipin stiffened TBM increased gene expression of pro-fibrotic cytokines and altered gene expression for N-cadherin, a proximal tubular epithelial specific cell-cell junction marker. This work introduces a new cell culture model for cell-basement membrane mechanobiology studies that utilizes in vivo-derived basement membrane. We also demonstrate that TBM stiffening affects tubular epithelial cell function through altered gene expression of cell-specific differentiation markers and induced increased expression of pro-fibrotic growth factors.

A novel approach for the endothelialization of xenogeneic decellularized vascular tissues by human cells utilizing surface modification and dynamic culture

Decellularized xenogeneic vascular grafts can be used in revascularization surgeries. We have developed decellularization methods using high hydrostatic pressure (HHP), which preserves the extracellular structure. Here, we attempted ex vivo endothelialization of HHP-decellularized xenogeneic tissues using human endothelial cells (ECs) to prevent clot formation against human blood. Slices of porcine aortic endothelium were decellularized using HHP and coated with gelatin. Human umbilical vein ECs were directly seeded and cultured under dynamic flow or static conditions for 14 days. Dynamic flow cultures tend to demonstrate higher cell coverage. We then coated the tissues with the E8 fragment of human laminin-411 (hL411), which has high affinity for ECs, and found that Dynamic/hL411showed high area coverage, almost reaching 100% (Dynamic/Gelatin vs Dynamic/hL411; 58.7 ± 11.4 vs 97.5 ± 1.9%, P = 0.0017). Immunostaining revealed sufficient endothelial cell coverage as a single cell layer in Dynamic/hL411. A clot formation assay using human whole blood showed low clot formation in Dynamic/hL411, almost similar to that in the negative control, polytetrafluoroethylene. Surface modification of HHP-decellularized xenogeneic endothelial tissues combined with dynamic culture achieved sufficient ex vivo endothelialization along with prevention of clot formation, indicating their potential for clinical use as vascular grafts in the future.

Design of hourglass-lattice metastructure with near-zero thermal expansion using structural optimization method

Zero thermal expansion (ZTE) is strongly demanded for critical components of spacecraft structures exposed to extreme temperature variations in space. In this paper, a lattice metastructure with near-ZTE is numerically achieved by structural optimization method. A sandwich panel with one single layer of hourglass-type lattice cells with variable cross-sectional and bi-material beams are assembled to produce either positive or negative thermal expansion (PTE or NTE). Then, a multilayered construction of metastructures is proposed by the combination of both PTE and NTE layers to realize the overall behaviors of near-zero deformation. The structural optimization with the gradient-based algorithm is utilized to obtain an extreme low value of coefficient of thermal expansion (∼10-9 /K). The shape constraint is adopted to regulate the thermal deformation of contact surfaces of each layer. Furthermore, bi-material prototypes with nylon and short carbon fiber reinforced composites are fabricated and tested, and the results demonstrate a NTE value of −6.32 × 10-5 /K, PTE value of + 1.28 × 10-4 /K, and ZTE value of 2.53 × 10-6 /K. This agrees well with the optimization result. Overall, the proposed hourglass-lattice metastructure with a high thermal stability is a promising solution to enable large-scale space structures such as space telescope, antenna, and radar, etc.

Cystic angiomatosis in children: clinical experience and review of literature

Cystic angiomatosis is a rare benign disease manifesting as multiple lytic and sclerotic bone lesions, described as the proliferation of vascular and lymphatic channels lined by a single layer of endothelial cells. However, the potential pathogenetic mechanism of the disease still remains unknown. Here, we reported a case of cystic angiomatosis with multifocal bone lesion evaluated by whole exome sequencing. In this presentation, we reported a case of an 11-year-old boy with pain in his chest. Computed tomography (CT) revealed the multiple lytic of the bone in the ribs, clavicle, vertebra thoracalis, skull, mandibula, shoulder blade, etc. The blood test showed ALP to be 393U/L and VEGF to be 287.26 pg/ml. The patient was performed with an open biopsy in the ribs and was diagnosed with cystic angiomatosis. Besides, the whole exome sequencing reported the single-nucleotide substitutions in the coding region of BRIP1, CHEK2, GRM4, and MUC16. Then, the upregulated genes involved CASC15, CENPF, ABCA13, ALK, BLM, and FGFR3. In this article, we report a rare case of cystic angiomatosis in a child with abnormal VEGF and ALP reported by peripheral blood examination. The whole exome sequencing could provide the reference for the potential molecular mechanism in the diagnosis and treatment of cystic angiomatosis.

Corneal endothelium – Specular microscopy

AbstractCorneal endothelium is a single layer of hexagonal cells covering the inner surface of the cornea and maintains the corneal deturgescence by continuous pumping of water and ions from the stroma to the aqueous humour. This mechanism is necessary to preserve the transparency of the cornea. The corneal endothelium is derived from the neural crest. Endothelial cells are arranged in a mosaic pattern of mostly hexagonal shaped cells. The cells can be characterized by different size (polymegathism) and different shape (pleomorphism). These parameters may reflect a compromised endothelial functional status.The corneal endothelial cell layer does not capable for regeneration, a constant and age‐dependent loss of corneal cells present during lifetime. Extensive injury of corneal endothelium leads to permanent corneal swelling and bullous alterations on the corneal surface. If left untreated and in chronic cases inflammation, later on permanent scarring may occur leads to loss of corneal transparency and finally decreased vision.Generally, a healthy cornea has about 1500–3500 endothelial cells/mm2 in the central area (age 40–75). The gold standard for the evaluation of endothelial cell density is endothelial microscopy, based on specular reflection, when light is reflected from the interfaces of materials with different indices of refraction. In addition, in vivo confocal microscope can also be used to visualize the endothelium, along with other tissue components of the cornea.Different forms of corneal damage by inflammatory processes or by mechanical trauma following intraocular surgery can lead to additional endothelial cell loss. Evaluation and proper examination of corneal endothelium and endothelial cell density is important and has clinical relevance in aging, corneal preservation, phacoemulsification, and post‐keratoplasty corneas.