European Journal of Histochemistry最新文献

This study investigates the expression and biological role of caudal homologous transcription factor 2 (CDX2) in duodenal carcinoma. Paraffin-embedded samples from 40 duodenal carcinoma cases were analyzed using immunohistochemistry on a tissue microarray to assess CDX2 expression and its prognostic significance. CDX2 overexpression plasmids and CDX2-siRNA were transfected into colon and duodenal cancer cells. Transfection efficiency was confirmed by RT-PCR and Western blotting. Cell proliferation was assessed using CCK8 assay, migration via scratch assay, and cell cycle and apoptosis by flow cytometry. CDX2 staining was primarily nuclear, with a positive rate of 65% in duodenal carcinoma tissues, significantly lower than in adjacent non-tumor tissues (p<0.05). CDX2-positive patients had better prognoses than negative patients (p<0.05). Reduced CDX2 expression significantly enhanced the proliferation of CaCO2 and HuTu-80 cells (p<0.001), whereas CDX2 overexpression suppressed proliferation (p<0.001). CDX2 knockdown increased migration, while its overexpression reduced migration (p<0.05). CDX2 overexpression led to a significant increase in G0/G1 phase cells and a decrease in S phase cells (p<0.05), whereas knockdown reduced G0/G1 phase cells and increased S and G2/M phase cells (p<0.05). Apoptosis was significantly increased following CDX2 overexpression (p<0.001) and decreased after CDX2 knockdown (p<0.001). CDX2 expression is downregulated or lost in duodenal carcinoma, acting as a tumor suppressor. CDX2 may serve as a crucial biomarker for predicting the onset, progression, and treatment of duodenal carcinoma.

The study examines the utility of AMACR, ERG, and AR immunostains in diagnosing prostatic adenocarcinoma (PCa) and assessing prognosis in comparison to the Gleason score and new WHO grading groups. Seventeen PCa biopsies and five benign prostatic hyperplasia (BPH) biopsies were analyzed. Immunoreactivity, scored from 1 to 3 based on percentage of positive cells and intensity of expression, was assessed, revealing 76.47% positivity for AMACR, 35.29% for ERG, and 94.12% for AR in PCa cases, with variable scores and intensity among markers and grade groups. AMACR sensitivity and ERG specificity were noted. Higher-grade PCa exhibited increased positivity for both markers, indicating prognostic significance. In BPH cases, AMACR showed positivity in 2 cases, ERG in 1, and AR in all cases, albeit with lower expression. Differential expression was observed among immunomarkers and grade groups of malignancy. AMACR and ERG stains serve as sensitive and specific markers for PCa diagnosis and prognosis. Their increasing positivity with higher-grade groups underscores prognostic value. These findings highlight the importance of immunostains in refining PCa diagnosis and prognostication.

In the present study, the expression of S100β was examined in the mouse cochlea from embryonic day 17 (E17) to postnatal day 32 (P32) using immunofluorescence, aiming to explore its possible role in auditory system. At E17, S100β expression was not detected, except in the external cochlear wall. Starting at E18.5, S100β staining appeared in the organ of Corti and the stria vascularis. In the E18.5 and P1 organ of Corti, S100β was confined to the developing pillar cells. By P6, cytoplasmic staining of S100β was evident in the inner and outer pillar cells, forming the tunnel of Corti. Additionally, S100β expression extended medially into the three rows of Deiter's cells, with labeling of their phalangeal processes. At P8, S100β continued to be expressed in the heads, bodies, and feet of the two pillar cells, as well as in the soma and phalangeal processes of the three rows of Deiter's cells. In the lateral wall of the P8 cochlea, S100β was expressed not only in the stria vascularis but also in the spiral ligament. Between P10 and P12, S100β expression was maintained in the Deiter's cells and pillar cells of the organ of Corti, as well as in the lateral wall, and spiral limbus. From P14 onwards, S100β expression ceased in the stria vascularis, though it persisted in the spiral ligament and spiral limbus into adulthood. Within the P14 and P21 organ of Corti, S100β remained in the Deiter's and pillar cells. S100β immunostaining was not observed in the phalangeal processes of Deiter's cells but was specifically present in the Deiter's cell cups at P21. In the adult cochlea (P28 and P32), S100β expression declined in both Deiter's and pillar cells. The dynamic spatiotemporal changes in S100β expression during cochlear ontogeny suggest its role in cochlear development and hearing function.

This study aimed to evaluate the therapeutic efficacy of camellia oil on 2,4-dinitrochlorobenzene (DNCB)-induced atopic dermatitis (AD) in mice, as well as its effect on the expression of skin-barrier-related proteins. A mouse model of AD was created via topical application of DNCB; subsequently, the animals were randomly divided into four groups: the blank control (Control), model (Model), moisturizing cream (Moisturizer), and camellia oil (Camellia) groups. The Camellia group received camellia oil, whereas the Moisturizer group was treated with moisturizing cream, as a positive control. Skin lesions, ear and back tissue morphology, and the serum levels of IgE, IL-4, and IFN-γ were analyzed. Compared with the Control group, AD mice exhibited erythema, papules, dryness, peeling, and significantly higher serum IgE and IL-4 levels. Compared with the Model group, treatment with camellia oil and moisturizing cream considerably reduced skin inflammation, ear thickness, and scratching frequency. A histopathological analysis revealed that camellia oil reduced inflammatory-cell infiltration and edema in the AD-affected skin. Furthermore, camellia oil upregulated filaggrin (FLG), thus aiding in skin-barrier repair. These findings suggest that camellia oil significantly improves AD symptoms, enhances FLG expression, and restores the damaged skin barrier in AD mouse models.

Accurate differentiation of homologous proteins that share high sequence identity remains a significant challenge in biomedical research, as conventional antibodies often lack sufficient specificity, leading to potential misinterpretations. This issue is particularly evident in the study of hexokinases, a family of isoenzymes that catalyze the first step of glycolysis by phosphorylating glucose. Beyond their canonical metabolic roles, hexokinases play critical non-glycolytic functions, especially in cancer biology. However, their unique tissue distributions and context-dependent roles are often obscured by the overlapping specificities of commercially available antibodies, which can produce misleading results. In this study, we rigorously evaluated a panel of antibodies targeting hexokinase isoenzyme 3 (HK3), highlighting the widespread issue of cross-reactivity and insufficient validation. Through this process, we identified and validated a highly specific antibody for HK3, demonstrating its reliability in western blot and immunohistochemistry applications. Using this validated tool, we reveal the distinct localization of HK3 in myeloid cell populations, providing new insights into its potential functional roles in these cells. This work addresses a critical gap in antibody specificity and establishes HK3 as a uniquely expressed gene in myeloid and immune cells and is absent in other cell types under basal conditions. Providing a foundation for future investigations into its context-dependent functions.

Muscle spindles are skeletal muscle sensory receptors composed of intrafusal fibres, partially encapsulated by connective tissue capsule. This capsule encloses the central A and B regions while leaving the distal C region extracapsular. Several past studies in rat have shown that muscle spindles typically contain a single bag1 fibre, a single bag2 fibre, and two smaller chain fibres. Intrafusal fibres co-express multiple myosin heavy chain (MyHC) isoforms: -slow or -1, -slow-tonic, -α, -2a, -2b, -embryonic, and -neonatal. While MyHC-2x was previously thought absent, the recently discovered MyHC-15 isoform has been identified in the C region of rat bag fibres. Using antibodies specific for nine MyHC isoforms and analyzing four different rat skeletal muscles-soleus, extensor digitorum longus, and the lateral and medial heads of gastrocnemius-we aimed to further characterize the co-expression pattern of MyHC-15 with other known isoforms and to determine whether MyHC-2x is expressed in rat intrafusal fibres. While rodents are widely used as animal models in skeletal muscle research, notable species-specific differences in MyHC isoform expression exist. Our findings revealed that MyHC-15 expression in rat intrafusal fibres is less abundant than in human fibres. MyHC-15 was primarily observed in bag fibres but was not detected in the C region, contrary to previous reports in both rat and human. We confirmed the absence of MyHC-2x in rat intrafusal fibres. Similarly, MyHC-embryonic and -neonatal were not detected in the analyzed spindles, suggesting that previously used antibodies may have cross-reacted with MyHC-2a and -2b. While our results partially corroborate previous extensive studies, discrepancies suggest that MyHC expression in intrafusal fibres varies not only along the fibre length but also across muscles.

This corrects the article published in European Journal of Histochemistry 2024;68:4140 doi: 10.4081/ejh.2024.4140 IF: 2.1 Q4 B4 IF: 2.1 Q4 B4.

Carbonic anhydrase (CA) has been localized to many structures involved in ion transport including the acini and ducts of the major (parotid, sublingual and submandibular) salivary glands of humans and rodents. It also has been localized by enzyme histochemistry and by immunohistochemistry for CA isoenzyme VI (CA VI) to the acini and ducts of rat serous lingual glands of von Ebner. The purpose of this study was to explore the intracellular distribution by cell type of three CA isoenzymes in these glands. Immunohistochemistry was undertaken with antibodies to human CAs I, II and VI in paraffin sections of rat tongues that had been fixed in Helly's fluid. The density of the reaction product was scored as 0 (none) to 5 (strongest). Reactions in the acini with CA I and II antibodies were weak luminally to moderate basally and generally moderate, respectively, moderate in the intercalated ducts, and moderate basally to strong luminally in the excretory ducts. Weak to moderate CA VI reactions occurred in the acini and ducts. The stronger luminal reactions to CAs I and II in the excretory ducts suggest that they contribute to pH regulation in the saliva of von Ebner's glands via HCO3- transport.

Intestinal barrier damage causes an imbalance in the intestinal flora and microbial environment, promoting a variety of gastrointestinal diseases. This study aimed to explore the mechanism by which adipose-derived stem cells (ADSCs) repair intestinal barrier damage. The human colon adenocarcinoma cell line Caco-2 and rats were treated with lipopolysaccharide (LPS) to establish in vitro and in vivo models, respectively, of intestinal barrier damage. The expression of inflammatory cytokines (TNF-α, HMGB1, IL-1β and IL-6), antioxidant enzymes (iNOS, SOD and CAT), and oxidative products (MDA and 8-iso-PGF2α) was detected using ELISA kits and related reagent kits. Apoptosis-related proteins (Bcl-2, Bax, Caspase-3 and Caspase-9), tight junction proteins (ZO-1, Occludin, E-cadherin, and Claudin-1) and p38 MAPK pathway-associated protein were detected by Western blotting. In addition, cell viability and apoptosis was determined by a CCK-8 kit and flow cytometry, respectively. Cell permeability was assayed by the transepithelial electrical resistance value and FITC-dextran concentration. The homing effect of ADSCs was detected by fluorescence labeling, and intestinal barrier tissue was observed by HE staining. After ADSC treatment, the level of phosphorylated p38 MAPK protein decreased, the expression of inflammatory factors, oxidative stress and cell apoptosis decreased, the expression of tight junction proteins increased, and cell permeability decreased in Caco-2 cells stimulated with LPS. In rats, ADSCs are directionally recruited to damaged intestinal tissue. ADSCs significantly decreased the levels of D-lactate, diamine oxidase (DAO) and FITC-dextran induced by LPS. ADSCs promoted tight junction proteins and inhibited oxidative stress in intestinal tissue. These effects were reversed after the use of a p38 MAPK activator. ADSCs can be directionally recruited to intestinal tissue, upregulate tight junction proteins, and reduce apoptosis and oxidative stress by inhibiting the p38MAPK signaling pathway. This study provides novel insights into the treatment of intestinal injury.

Colorectal cancer (CRC) is a major public health concern and identifying prognostic molecular biomarkers can help stratify patients based on risk profiles, thus enabling personalized medicine. Epitranscriptomic modifications play a relevant role in controlling gene expression, N6-methyladenosine (m6A) regulators play crucial roles in cancer progression, but their clinical significance in CRC cancer has thus far not been elucidated. Thus, we aimed to examine by immunohistochemical techniques and RT-qPCR, protein levels and RNAs expression of m6A writers (METTL3, WTAP) and eraser (FTO) in a cohort of 10 patients affected by CRC. The patients were followed for 5 years and values of METTL3, WTAP and FTO RNAs in alive vs dead patients were compared. Proteins expression and RNAs expression had a different trend, METTL3, WTAP and FTO proteins' expression showed an increasing trend from non-cancerous adjacent (N) tissue vs carcinoma (CA) tissue G1 stage, and then a decreasing trend from G1 to G2 and G3 stages. The most marked increase was observed in WTAP that, from a 40% of protein expression positivity in N tissue raised to the 81% of positivity in G1 stage K tissue. RNAs expression of METTL3, WTAP and FTO genes in N tissue vs G1 stage CA tissue was significantly different, the analysis and comparison of RNAs values in patient alive after 5 years (0.58±0.04) vs patients dead after 5 years (1.69±0.29) showed that only WTAP values resulted significantly high in dead patients. The fact that WTAP protein expression levels lower while WTAP RNA expression remains high, lets us hypothesize a sort of inhibition of protein expression, but further studies are needed to clarify the mechanism. Although the results suggest a relationship between biological meaning and prognostic utility of WTAP, this prognostic utility must be confirmed by further studies on a larger sample.