Abnormal Epithelial Differentiation Research Articles

Reduced Proliferative Capacity and Defense against Staphylococcus aureus in Human Nasal Mucosal Epithelium Lacking ZNF365

Introduction: Chronic rhinosinusitis with nasal polyps (CRSwNP) is a common chronic inflammatory disease of the nose characterized by barrier disruption and environmental susceptibility, and the deletion of ZNF365 may be a factor inducing these manifestations. However, there is no study on the mechanism of action between CRSwNP and ZNF365. Therefore, this study focuses on the effect of the zinc finger protein ZNF365 on the proliferation of nasal mucosal epithelial cells and their defense against Staphylococcus aureus (S. aureus). Methods: Immunohistochemistry and Western blot were applied to verify the changes of ZNF365 expression in nasal polyp tissues and control tissues, as well as in primary epithelial cells. ZNF365 was knocked down in human nasal mucosa epithelial cell line (HNEpc), and the proliferation, migration, and transdifferentiation of epithelium were observed by immunofluorescence, QPCR, CCK8, and cell scratch assay. The changes of mesenchymal markers and TLR4-MAPK-NF-κB pathway were also observed after the addition of S. aureus. Results: ZNF365 expression was reduced in NP tissues and primary nasal mucosal epithelial cells compared to controls. Knockdown of ZNF365 in HNEpc resulted in decreased proliferation and migration ability of epithelial cells and abnormal epithelial differentiation (decreased expression of tight junction proteins). S. aureus stimulation further inhibited epithelial cell proliferation and migration, while elevated markers of epithelial-mesenchymal transition and inflammatory responses occurred. Conclusion: ZNF365 is instrumental in maintaining the proliferative capacity of nasal mucosal epithelial cells and defending against the invasion of S. aureus. The findings suggest that ZNF365 may participate in the development of CRSwNP.

HS 3D-SeboSkin Model Enables the Preclinical Exploration of Therapeutic Candidates for Hidradenitis Suppurativa/Acne Inversa

Despite the rapid development in hidradenitis suppurativa (HS) research, the immediate introduction of potent therapeutic compounds in clinical trials and the lack of definitive outcome measures have led to the discontinuation of potential therapeutic compound studies. HS is a solely human disease, and therefore, the search for preclinical human models has been given priority. The 3D-SeboSkin model, a co-culture of human skin explants with human SZ95 sebocytes as a feeder layer, has been shown to prevent the rapid degeneration of human skin in culture and has been validated for HS preclinical studies. In this work, the HS 3D-SeboSkin model has been employed to characterize cellular and molecular effects of the EMA- and FDA-approved biologic adalimumab. Adalimumab, a tumor necrosis factor-α inhibitor, was shown to target inflammatory cells present in HS lesions, inducing a prominent anti-inflammatory response and contributing to tissue regeneration through a wound healing mechanism. Adalimumab inhibited the lesional tissue expression of TNF-α, IL-3, IL-15, and MCP-3 and downregulated the secretion of IL-1α, IL-5, RANTES, MCP-2, TNF-α, TNF-β, TGF-β, and IFN-γ. In contrast, IL-6 was stimulated. The compound failed to modify abnormal epithelial cell differentiation present in the HS lesions. Patients with Hurley stage II lesions exhibited stronger expression of autophagy proteins in perilesional than in lesional skin. Adalimumab modified the levels of the pro-apoptotic proteins LC3A, LC3B, and p62 in an individual, patient-dependent manner. Finally, adalimumab did not modify the NFκB signal proteins in SZ95 sebocytes and NHK-19 keratinocytes, used to study this specific pathway. The administration of the validated HS 3D-SeboSkin model in ex vivo studies prior to clinical trials could elucidate the individual pathogenetic targets of therapeutic candidates and, therefore, increase the success rates of clinical studies, minimizing HS drug development costs.

Loss of basigin expression in uterine cells leads to subfertility in female mice†.

Basigin (BSG) is a transmembrane glycoprotein involved in cell proliferation, angiogenesis, and tissue remodeling. BSG has been shown to be essential for male and female reproduction although little is known about its role in normal uterine function. To study the potential function of BSG in the female reproductive tract, we generated mice with conditional knockout of Bsg in uterine cells using progesterone receptor-Cre and hypothesized that BSG is required for normal pregnancy in mice. Fertility study data showed that the conditional knockout mice had significantly reduced fertility compared to controls. Ovarian function of the conditional knockout mice appeared normal with no difference in the number of superovulated oocytes collected or in serum progesterone levels between the conditional knockout and the control mice. Uterine tissues collected at various times of gestation showed increased abnormalities in implantation, decidualization, placentation, and parturition in the conditional knockout mice. Uterine cross sections on Day 5 of pregnancy showed implantation failure and abnormal uterine epithelial differentiation in a large proportion of the conditional knockout mice. There was a compromised decidual response to artificial decidualization stimuli and decreased mRNA and protein levels for decidualization genes in the uteri of the conditional knockout mice. We also observed altered protein expression of monocarboxylate transporter 1 (MCT1), as well as impaired angiogenesis in the conditional knockout uteri compared to the controls. These results support that BSG is required for successful pregnancy through its functions in implantation and decidualization.

Epithelial to mesenchymal transition-related proteins ZEB1, β-catenin, and β-tubulin-III in idiopathic pulmonary fibrosis

Epithelial to mesenchymal transition has been suggested as a relevant contributor to pulmonary fibrosis, but how and where this complex process is triggered in idiopathic pulmonary fibrosis is not fully understood. Beta-tubulin-III (Tubβ3), ZEB1, and β-catenin are partially under the negative control of miR-200, a family of micro-RNAs playing a major role in epithelial to mesenchymal transition, that are reduced in experimental lung fibrosis and idiopathic pulmonary fibrosis. We wonder whether in situ expression of these proteins is increased in idiopathic pulmonary fibrosis, to better understand the significance of miR-200 feedback loop and epithelial to mesenchymal transition. We investigated the immunohistochemical and immunofluorescent expression and precise location of ZEB1, Tubβ3, and β-catenin in tissue samples from 34 idiopathic pulmonary fibrosis cases and 21 controls (5 normal lungs and 16 other interstitial lung diseases). In 100% idiopathic pulmonary fibrosis samples, the three proteins were concurrently expressed in fibroblastic foci, as well in damaged epithelial cells overlying these lesions and in pericytes within neo-angiogenesis areas. These results were also confirmed by immunofluorescence assay. In controls the abnormal expression of the three proteins was absent or limited. This is the first study that relates concurrent expression of Tubβ3, ZEB1, and β-catenin to abnormal epithelial and myofibroblast differentiation in idiopathic pulmonary fibrosis, providing indirect but robust evidence of miR-200 deregulation and epithelial to mesenchymal transition activation in idiopathic pulmonary fibrosis. The abnormal expression and localization of these proteins in bronchiolar fibro-proliferative lesions are unique for idiopathic pulmonary fibrosis, and might represent a disease-specific marker in challenging lung biopsies.

Gpr177 regulates pulmonary vasculature development

Establishment of the functional pulmonary vasculature requires intimate interaction between the epithelium and mesenchyme. Previous genetic studies have led to inconsistent conclusions about the contribution of epithelial Wnts to pulmonary vasculature development. This discrepancy is possibly due to the functional redundancy among different Wnts. Here, we use Shh-Cre to conditionally delete Gpr177 (the mouse ortholog of Drosophila Wntless, Wls), a chaperon protein important for the sorting and secretion of Wnt proteins. Deletion of epithelial Gpr177 reduces Wnt signaling activity in both the epithelium and mesenchyme, resulting in severe hemorrhage and abnormal vasculature, accompanied by branching defects and abnormal epithelial differentiation. We then used multiple mouse models to demonstrate that Wnt/β-catenin signaling is not only required for the proliferation and differentiation of mesenchyme, but also is important for the maintenance of smooth muscle cells through the regulation of the transcription factor Kruppel-like factor 2 (Klf2). Together, our studies define a novel mechanism by which epithelial Wnts regulate the normal development and maintenance of pulmonary vasculature. These findings provide insight into the pathobiology of congenital lung diseases, such as alveolar capillary dysplasia (ACD), that have abnormal alveolar development and dysmorphic pulmonary vasculature.

Abnormal Epithelial Differentiation and Tear Film Alteration in Pinguecula

To investigate the differentiation of conjunctival epithelium and tear film function in pingueculae. Twelve patients (12 eyes) who underwent simple excision for pingueculae were enrolled in the study. Immunostaining for K14, K19, K10, MUC5AC, Pax6, P63, Ki67, and K16 was performed on the pinguecula epithelium and normal conjunctival epithelium. Schirmer I test results and tear film break-up time (TFBUT) were evaluated just before and 1 month after surgery. Abnormal epidermal differentiation of pinguecula tissue, as evidenced by a decline in or absence of Pax6 expression, was accompanied by a decline in or absence of K19 keratin and MUC5AC, and an increase in K10 and K14 keratin. Furthermore, the pinguecula epithelium was actively proliferating, as evidenced by positive expression of Ki67, P63, and K16 keratin. The Schirmer I test results did not indicate any significant differences before and after surgery. However, the TFBUT was significantly prolonged 1 month after surgery compared with before surgery. Pinguecula is a condition of abnormal epithelial differentiation. It is characterized by squamous proliferation and metaplasia, resulting in instability of tear film with normal basic tear secretion.

Abnormal differentiation, hyperplasia and embryonic/perinatal lethality in BK5-T/t transgenic mice

The cell-of-origin has a great impact on the types of tumors that develop and the stem/progenitor cells have long been considered main targets of malignant transformation. The SV40 (SV40—Simian Virus 40) large T and small t antigens (T/t), have been targeted to multiple-differentiated cellular compartments in transgenic mice. In most of these studies, transgenic animals develop tumors without apparent defects in animal development. In this study, we used the bovine keratin 5 (BK5) promoter to target the T/t antigens to stem/progenitor cell-containing cytokeratin 5 (CK5) cellular compartment. A transgene construct, BK5-T/t, was made and microinjected into the male pronucleus of FVB/N mouse oocytes. After implanting ∼1700 embryos, only 7 transgenics were obtained, including 4 embryos (E9.5, E13, E15, and E20) and 3 postnatal animals, which died at P1, P2, and P18, respectively. Immunohistological analysis revealed aberrant differentiation and prominent hyperplasia in several transgenic CK5 tissues, especially the upper digestive organs (tongue, oral mucosa, esophagus, and forestomach) and epidermis, the latter of which also showed focal dysplasia. Altogether, these results indicate that constitutive expression of the T/t antigens in CK5 cellular compartment results in abnormal epithelial differentiation and leads to embryonic/perinatal animal lethality.

Genetic Ablation of αv Integrins in Epithelial Cells of the Eyelid Skin and Conjunctiva Leads to Squamous Cell Carcinoma

Integrin-mediated cell adhesion and signaling events are essential for the proper development and homeostasis of most epithelial tissues. Dysregulation of integrin expression and function can cause abnormal epithelial cell proliferation and/or differentiation, contributing to the pathogenesis of malignant epithelial cancers. Here we report on the use of a conditional knockout strategy exploiting the Cre/Lox technology to study the in vivo functions of alphav integrins during epithelial cell proliferation and differentiation. We show that genetic ablation of alphav integrin expression in basal epithelial cells of the eyelid skin and conjunctiva causes the formation of tumors that are strikingly similar to the malignant epithelial cancer, squamous cell carcinoma. These data suggest a mechanism whereby alphav integrins normally suppress epithelial cell proliferation, likely via adhesion to ECM ligands, as well as by the modulation of intracellular signaling cascades. We propose that alphav gene deletion eliminates normal integrin-mediated growth suppression, ultimately leading to cellular transformation and tumorigenesis. Hence, these studies reveal a novel tumor suppressor-like function of alphav integrins and provide a genetically tractable mouse model for studying the pathogenesis of squamous cell carcinoma and related cancers of epithelial origin, as well as to test and develop novel therapeutic compounds to treat or prevent squamous cell carcinoma of the skin.

Ocular Manifestations and Histologic Characteristics of Keratitis-Ichthyosis-Deafness (KID) Syndrome

Purpose: Keratitis-ichthyosis-deafness (KID) syndrome is a congenital ectodermal disorder presenting the triad of vascularizing keratitis, erythrokeratoderma, and sensorineural deafness. Ocular manifestations such as hyperkeratinization of the eyelids, loss of eyelashes, keratoconjunctivitis sicca, corneal erosions, ulceration, neovascularization, and scarring opacity may be absent or mild, but if present and severe, they can lead to major visual loss. We report a patient with KID syndrome with severe ocular manifestations and the histologic characteristics of a corneal lesion. Case summary: A 5-year-old boy was referred to the Ophthalmology Department for bilateral hyperkeratinization of eyelids, bare eyelashes, and corneal opacity. He showed hyperkeratotic skin lesions and sensorineural hearing loss. Molecular analysis showed a mutation in the GJB2 gene and confirmed the diagnosis of KID syndrome. Initial conservative treatment did not preserve ocular surface integrity, and instead it was maintained by surgical procedures including superficial lamellar keratectomy with amniotic membrane transplantation. The histologic characteristics of corneal lesions are abnormal epithelial differentiation, absence of connexin 26 expression, and partial destruction of the basement membrane.

HPV8 early genes modulate differentiation and cell cycle of primary human adult keratinocytes

Human papillomaviruses (HPV) have been associated with the development of non-melanoma skin cancer (NMSC) but the molecular mechanisms of the role of the virus in NMSC development are not clearly understood. Abnormal epithelial differentiation seen in malignant transformation of keratinocytes is associated with changes in keratin expression. The purpose of this study was to investigate the phenotype of primary human adult keratinocytes expressing early genes of HPV8 with specific reference to their differentiation and cell cycle profile to determine whether early genes of HPV8 lead to changes that are consistent with transformation. The expression of HPV8 early genes either individually or simultaneously caused distinct changes in the keratinocyte morphology and induced an abnormal keratin expression pattern, that included simple epithelial (K8, K18, K19), hyperproliferation-specific (K6, K16), basal-specific (K14, K15) and differentiation-specific (K1, K10) keratins. Our results indicate that expression of HPV8 early genes disrupts the normal keratin expression pattern in vitro. Expression of HPV8-E7 alone caused polyploidy that was associated with decreased expression of p21 and pRb. Expression of individual genes or in combination differentially influenced cell morphology and cell cycle distribution which might be important in HPV8-induced keratinocyte transformation.

Oral lichen planus treated with 13-cis-retinoic acid (isotretinoin): effects on the apoptotic process

The aim of the present study is to verify the efficacy of isotretinoin in oral lichen planus (OLP). In a double-blind study, ten patients with biopsy-proven OLP were treated for 4 months with 0.1% isotretinoin gel and another ten patients with placebo. At the end of the first period of observation, the patients who had been given the placebo were given isotretinoin for a further 4 months. A complete response was defined as the disappearance of the lesions as assessed by inspection, whereas a partial response was defined as a 50% or more reduction in the size of the lesions. All patients treated with isotretinoin showed a significant improvement of the oral lesions, whereas in the patients who were given the placebo, the size of the lesions remained the same. The patients who were given isotretinoin after the placebo showed a reduction in lesions. In total, there were ten complete and ten partial responses. Lesions were analysed histologically and immunohistochemically with antibodies against bcl-2 and Ki-67. Ki-67 and bcl-2 have statistical significant increased values from before to after treatment, whereas apoptotic bodies decreased one. All these facts could have contributed to the partial or complete regression of OLP lesions. The increase in Ki-67 positive cells show that the epithelium requires for enhanced proliferation and healing. The present results revealed a disturbed cell death programme in OLP that could underline an abnormal epithelial differentiation. The results of this pilot study show that the topical use of isotretinoin is effective in treating OLP.

Differential expression of matrix metalloproteinases and interleukin‐8 during regeneration of human airway epithelium in vivo

In many airway diseases, the airway epithelium is severely damaged and has to regenerate rapidly to restore its function. The regeneration process involves chronological steps of epithelial cell migration, proliferation, stratification, and differentiation. The present study has used an in vivo humanized airway xenograft model in nude mice that mimics the regeneration dynamics of human airway epithelium after severe injury, and human-specific molecular tools, to study the expression profiles of epithelial matrix metalloproteinases (MMPs)-7 and -9, of tissue inhibitor of matrix metalloproteinase-1 (TIMP-1), and of the pro-inflammatory cytokine interleukin-8 (IL-8) during the different steps of human airway epithelium regeneration. It was found that during the cell migration and proliferation steps, airway epithelial cells expressed IL-8 at a high level, whereas airway epithelial pseudo-stratification and surface airway epithelial differentiation were associated with increased expression of MMPs and a progressive decrease in IL-8. Interestingly, immunohistochemical analysis revealed exclusive expression of MMPs at the apical part of the well-differentiated regenerated airway epithelium, and incubation of the regenerating epithelial cells with MMP inhibitors led to abnormal epithelial differentiation. These data provide new insight into the temporal expression of MMPs and IL-8 during the regeneration of airway epithelium and demonstrate the involvement of these factors during the different steps that lead to restoration of a well-differentiated and functional airway epithelium.

Developmental and cellular factors underlying corneal epithelial dysgenesis in the Pax6+/− mouse model of aniridia

Heterozygosity for a PAX6 deficiency ( PAX6 +/− ) results in low levels of the PAX6 transcription factor and causes aniridia. Corneal changes in aniridia-related keratopathy (ARK) include peripheral pannus and epithelial abnormalities, which eventually result in corneal opacity and contribute to visual loss. The corneal abnormalities of Pax6 +/− mice provide an excellent model for the corneal changes seen in PAX6 +/− humans. The aim of the present study was to investigate the contributions of different factors (including altered cell proliferation, abnormal epithelial differentiation and incursion of the conjunctival epithelium) that may underlie the pathogenesis of the corneal changes caused by low levels of Pax6 in heterozygous Pax6 +/Sey-Neu ( Pax6 +/− ) mice. BrdU incorporation showed enhanced proliferation of Pax6 +/− corneal epithelium compared to wild-type controls and analysis of p63 (a marker of high proliferative potential) revealed a slight increase in frequency of p63-positive basal corneal epithelial cells in Pax6 +/− mice. Immunohistochemical investigation of K12 (a Pax6-regulated marker of corneal epithelial differentiation) in 2–52-week-old mice showed that K12 expression was delayed and down-regulated in the Pax6 +/− corneal epithelium, implying that differentiation of the Pax6 +/− corneal epithelium was delayed and abnormal. Goblet cells were identified within the peripheral corneal epithelium of the Pax6 +/− eyes, but some were surrounded by cells expressing K12, suggesting they may have arisen in situ in the corneal epithelium. These findings suggest that low levels of Pax6 may be directly responsible for failure or delay of proper differentiation of the corneal epithelial cells, but the proliferative component of the mutant epithelium is probably not impaired. This abnormal differentiation suggests that ARK is not entirely attributable to a limbal stem cell deficiency.

Modulation of abnormal colonic epithelial cell proliferation and differentiation by low-fat dairy foods

Holt, Peter R.; Atillasoy, Evren O.; Gilman, Jody; Guss, Janet; Moss, Steven F.; Newmark, Harold; Fan, Kunhua; Yang, Kan; Lipkin, Martin

Restriction of Nutrition In Utero Selectively Inhibits Gastrointestinal Growth in Fetal Sheep , ,

This study examined the effects of reduced nutrition on fetal growth over the first half of gestation. Reduced nutrition was achieved by a combination of reduced maternal food intake and carunclectomy, a procedure which restricts the development of the placenta. There were no major effects of restriction on fetal body, tissue or organ growth, except for the gastrointestinal tract (GIT). Total GIT weight was lower in restricted fetuses than in controls. More specifically, it was growth of the small and large intestine which was compromised. Small intestinal weight was significantly lower, both in absolute terms and relative to body weight. The intestinal diameter and mucosal area were significantly lower in both small and large intestine of restricted fetuses. Maturation of enterocytes was also delayed in nutrient-restricted fetuses. In addition, there were focal lesions of the brush border present, indicating abnormal epithelial differentiation. By term, in growth-retarded fetuses, growth deficits in many organs were present, including the GIT. The present study suggests that GIT growth deficits may have a long-term etiology, including at their onset, abnormal cellular differentiation. These results could explain why GIT function in intrauterine growth-retarded infants is more likely to be compromised than in premature or term infants.

Tretinoin therapy in photoageing: historical perspective

Tretinoin was shown in the late 1960s to be useful for the treatment of disorders associated with abnormal epithelial differentiation; however, because of irritation, retinoids were only slowly accepted. In the 1970s, evidence accumulated to show that topical tretinoin could modulate many of the abnormalities in the epidermis and dermis associated with photoageing. It has been shown in hairless mice that tretinoin can reverse dermal elastosis with the formation of new collagen and this has led to clinical trials being carried out in man. Randomized, controlled trials have shown that topical tretinoin is effective in the treatment of photoaged skin.

Growth factors in wound healing. Single and synergistic effects on partial thickness porcine skin wounds.

Several growth factors are potential mediators of wound healing, although their actual roles, interactions, and therapeutic use are not established. Six well-characterized human growth factors were chosen for detailed investigation by topical application to standardized skin wounds in swine: epidermal growth factor (EGF), transforming growth factors alpha and beta (TGF-alpha and TGF-beta), fibroblast growth factor (FGF), insulin-like growth factor-I (IGF-I), and platelet-derived growth factor (PDGF). When applied singly in doses up to 1,500 ng, only TGF-beta produced a marked tissue response, as demonstrated by an increase in the new connective tissue volume, the collagen content and maturity, and increased angiogenesis. However, TGF-beta enhanced inflammation and caused abnormal epithelial differentiation and decreased epithelial volume, the last reversed by addition of IGF-I. Recombinant PDGF-2 homodimer, if given in combination with recombinant IGF-I, caused a similar increase in the new connective tissue volume and collagen content and maturity, but without increased inflammation. In addition, this combination stimulated increased amounts of epithelium with normal differentiation. The synergy of PDGF-2 and IGF-I was optimal at a ratio of 2:1 by weight. Of the six individual factors and nine combinations tested, the combinations of PDGF-2 and IGF-I or PDGF-2 and TGF-alpha were the most potent stimulators of healing in the absence of increased inflammation.

Renal epithelial development in organotypic culture.

An in vitro model system for the experimental study of renal epithelial differentiation is described. Fetal murine metanephric tissue consisting of nephrogenic blastema and branched ureteric bud is isolated following 24-36 h of natural embryonic inductive interaction (13 +/- 0.4 days gestation) and cultured as an intact organ in a Trowell-type assembly. During 120 h of organ culture incubation in completely defined serum-free medium, advanced organotypic proximal tubular and glomerular epithelial differentiation proceed in the absence of vascularization, perfusion, and urine production. The system thus experimentally separates the processes of three-dimensional organ growth and post-induction renal epithelial differentiation from glomerular filtration, flow-related phenomena, endothelial or mesangial cell interactions, and the effects of growth factors or transport substrates present in mammalian serum or urine. Studies to date in the model system have defined the growth factor requirements of epithelial growth and differentiation and have demonstrated that specific hormonally induced alterations in tubular epithelial cell metabolism and function may lead to specific patterns of tubular maldevelopment. Whole organ metanephric organ culture is thus a valuable in vitro model system for future investigations into the complex processes of normal and abnormal renal epithelial differentiation.

Histologic changes associated with the topical use of isotretinoin on oral lichen planus

B 0th systemic and topical synthetic retinoids (vitamin A derivatives) have been used extensively in the treatment of keratotic skin diseases that feature abnormal epithelial cell proliferation and differentiation. Dramatic results have been reported following the treatment of psoriasis and acne with etretinate and isotretinoin (13~cis retinoic acid).‘-4 Because of the antikeratinizing and immunomodulating effects of retinoids,‘.4*s the use of these compounds has been extended to the treatment of patients with cutaneous and oral lichen planus. Systemic retinoids-etretinate, isotretinoin, and tretinoin (all-trans retinoic acid)-have produced clinically significant results in patients with oral lichen planus.‘-l3 In one study, however, only marginal improvement was noted in patients with erosive lichen planus. I4 Because of side effects, the investigators thought that treatment with systemic etretinate did not justify the results. Topical tretinoin has produced generally good results in patients with oral lichen planus, without the complications of cheilitis, dry skin, eievation of serum liver enzymes, and elevation of serum triglycerides and cholesterol that are commonly associated with systemic therapy. ‘. ‘w’ Until recently, studies using topical agents have been limited to the use of tretinoin. A related retinoid-isotretinoin-had been made available to us in a 0.1% gel for clinical testing and showed considerable efficacy in the treatment of oral lichen planus. I8 Atrophic and erosive lesions responded less dramatically than reticular lesions. This double-blind clinical trial spanned 8 weeks of twice-daily application of isotretinoin gel (Figs. 1

Intracellular keratins in normal and pathological bronchial mucosa. Immunocytochemical studies on biopsies and cell suspensions.

The distribution of intracellular keratins was investigated in normal bronchial epithelium and in several morphologically distinct forms of respiratory tract carcinomas. This study was performed with two different experimentally produced antisera against normal human stratum corneum keratin and against keratin protein of MW 67,000 dalton, using indirect immunofluorescence and immunoperoxidase methods on tissue sections and cell suspensions. In normal bronchial epithelium, the basal cells were strongly labelled by both antisera. The ciliated columnar cells appeared devoid of cytokeratins in tissue sections but were strongly labelled with both antisera in cell suspensions. The goblet cells remained negative in every case. In squamous metaplasia of the bronchus, all epithelial cells were unevenly stained with both antisera. Among tumours, only the squamous cell carcinomas were strongly labelled by both antisera. Primary lung adenocarcinoma appeared weakly positive, whereas metastatic lung carcinomas, undifferentiated lung carcinomas, oat cell tumours, carcinoid tumours were negative. The immunocytochemical determination of keratins appeared to be of value in the study of normal and abnormal epithelial differentiation, in the diagnosis of poorly differentiated carcinomas and in their distinction from metastatic tumours of the lung.