Identification and culture of functional salivary gland ductal epithelial cells.

Sialadenitis is a prevalent salivary gland disease resulting in decreased salivary flow rate. To date, little is known about the exact changes and mechanism of ductal cells in sialadenitis. This study aims to establish an efficient method to identify and isolate ductal cells, thereby facilitating further research on this specific cell type. Immunofluorescence for cytokeratin 13 and cytokeratin 19 was conducted in salivary glands to confirm their specificity as ductal cell markers. The dissected ducts were assessed through PCR and Western blot of cytokeratin 19 and digested by dispase and collagenase. The functionality of the isolated ductal cells was determined by measuring intracellular calcium. Cytokeratin 19 and cytokeratin 13 were expressed in all segments of human ducts. Cytokeratin 19 was limited to ducts excluding granular convoluted tubules in rat and mouse. The purities of the obtained ductal cells were approximately 98% in humans and 93% in rats. Furthermore, intracellular free calcium increased with time and concentration of carbachol treatment. Cytokeratin 19 serves as a dependable marker for identifying ductal cells in salivary glands, except for granular convoluted tubules. Moreover, we have successfully developed an efficient method for isolating ductal cells from salivary glands.

Switch to a raltegravir-based antiretroviral regimen in people with HIV and non-alcoholic fatty liver disease: A randomized controlled trial.

INTRODUCTION: The effect of antiretroviral therapy (ART), particularly integrase strand transfer inhibitors (INSTIs), on non-alcoholic fatty liver disease (NAFLD) in people with HIV remains unclear. We evaluated the effect of switching non-INSTI backbone antiretroviral medications to raltegravir on NAFLD and metabolic parameters. MATERIALS AND METHODS: This was a single-centre, phase IV, open-label, randomized controlled clinical trial. People living with HIV with NAFLD and undetectable viral load while receiving a non-INSTI were randomized 1:1 to the switch arm (raltegravir 400 mg twice daily) or the control arm (continuing ART regimens not containing INSTI). NAFLD was defined as hepatic steatosis by controlled attenuation parameter ≥238 dB/m in the absence of significant alcohol use and viral hepatitis co-infections. Cytokeratin 18 was used as a biomarker of non-alcoholic steatohepatitis. Changes over time in outcomes were quantified as standardized mean differences (SMDs), and a generalized linear mixed model was used to compare outcomes between study arms. RESULTS: A total of 31 people with HIV (mean age 54 years, 74% male) were randomized and followed for 24 months. Hepatic steatosis improved between baseline and end of follow-up in both the switch (SMD -43.4 dB/m) and the control arm (-26.6 dB/m); the difference between arms was not significant. At the end of follow-up, aspartate aminotransferase significantly decreased in the switch arm compared with the control arm (SMD -9.4 vs. 5.5 IU/L). No changes in cytokeratin 18, body mass index, or lipids were observed between study arms. DISCUSSION: Switching to a raltegravir-based regimen improved aspartate aminotransferase but seemed to have no effect on NAFLD, body weight, and lipids compared with remaining on any other ART.

Feasibility study on epithelial cells regeneration after excising the demucosalized ileum for ureteral reconstructive surgery.

This study aimed to investigate the regeneration of epithelial cells in the long-term observation of ureter reconstruction by excising the demucosalized ileum. First, 8 Beagle dogs were anesthetized and the abdominal cavity was inspected for abnormalities via an abdominal incision. The right kidney and ureter were subsequently separated, and the ureter was severed from its connection to the renal pelvis and bladder and ligated distally. The 10-15 cm of ileum was used to reconstruct the ureter. The biopsies of the proximal, middle, and distal reconstructed ureter (neo-ureter) were collected at the first, third, fifth, and sixth month postoperatively. The regeneration of ileal mucosa at the first, third, fifth, and sixth month was observed by hematoxylin-eosin (HE) staining and immunofluorescence staining cytokeratin 18 (CK18). HE staining results showed irregular cytoarchitecture, severe nuclear consolidation, and inflammatory infiltration in the proximal, middle, and distal neo-ureters of dogs at the first month after ureteral reconstruction. With longer follow-up, the injuries of the proximal, middle, and distal neo-ureters were alleviated at the third, fifth, and sixth month after surgery. The expression of CK18 was higher in the middle neo-ureters than that in the proximal and distal neo-ureters at different time points after ureteral reconstruction and decreased with time. In summary, the present study demonstrated that demucosalized ileum was feasible for ureteral reconstructive surgery with satisfying prognostic effects.

A Mouse Model for Conditional Expression of Activated ß-Catenin in Epidermal Keratinocytes.

We report the generation and characterization of the K5: CAT bigenic mouse in which the constitutively activated form of ß-catenin (ΔN89 ß-catenin) is conditionally expressed in cytokeratin-5 (K5) positive epidermal keratinocytes. Following short-term doxycycline intake during the telogen resting phase, the adult K5: CAT bigenic develops enlarged pilosebaceous units that expand deep into the dermis, an expansion usually observed during the anagen growth phase. Prolonged doxycycline treatment results in significant thickening and folding of the K5: CAT epidermis. During this persistent induction period, there is clear evidence of increased keratinocyte proliferation, particularly in the epidermal basal cell layer and the outer root sheath of the hair follicle. This unscheduled increase in cellular proliferation likely explains the decrease in hair density observed in the K5: CAT mouse following persistent doxycycline intake. Numerous hyperplastic endometrioid cysts, which display cornification toward their lumens, are also observed during this treatment period. Remarkably, de-induction of ΔN89 ß-catenin expression through doxycycline withdrawal results in a marked reversal of the skin phenotype, suggesting that these morphological changes are dependent on continued signaling by ß-catenin and/or its downstream molecular mediators. Joining a small group of mouse models for conditional ß-catenin signaling, our K5: CAT mouse model will be particularly useful in identifying those molecular mediators of ß-catenin that are responsible for initiating and maintaining these phenotypic responses in the K5: CAT skin. Such studies are predicted to shed more light on ß-catenin signaling in epidermal epithelial morphogenesis, hair follicle cycling, and hair growth pathologies.

Fusion expression, purification, and characterization of cytokeratin 19 fragments in E. coli for enhanced stability in diagnostic applications.

Cytokeratin 19 fragment (CYFRA21-1) serves as a crucial tumor marker in the context of lung cancer patients, playing a pivotal role as a calibrator in the realm of in vitro diagnostics. Nevertheless, during practical application, it has come to light that the recombinantly synthesized full-length CYFRA21-1 antigen exhibits suboptimal stability at the requisite concentration, while the utilization of natural antigens incurs a substantial cost. To address this issue, our investigation harnessed a strategic approach whereby the soluble fragment of cytokeratin 19 (Aa244-400) was integrated into the pET32a vector, subsequently being expressed within E. coli through a fusion with the TrxA protein. This process involved induction of protein expression through 0.2 mM IPTG at 16 °C for a duration of 16 h. After induction, the target protein was purified through Ni affinity and ion exchange chromatography. Subsequent characterization of the targeted protein was executed through the SEC-HPLC technique. The attained CYFRA21-1 antigen, as generated within this study, was effectively incorporated into a chemiluminescence-based in vitro diagnostic detection kit. The results indicate that the fusion protein exhibited commendable reactivity and stability, manifesting a deviation of less than 10 % following incubation at 37 °C for 7 days. Importantly, the production yield achieved a notable magnitude of 300 mg/L, thus rendering it a cost-effective and scalable alternative to natural antigens for clinical diagnostic applications.

Placental differences between severe fetal growth restriction and hypertensive disorders of pregnancy requiring early preterm delivery: morphometric analysis of the villous tree supported by artificial intelligence.

BACKGROUND: The great obstetrical syndromes of fetal growth restriction and hypertensive disorders of pregnancy can occur individually or be interrelated. Placental pathologic findings often overlap between these conditions, regardless of whether 1 or both diagnoses are present. Quantification of placental villous structures in each of these settings may identify distinct differences in developmental pathways. OBJECTIVE: This study aimed to determine how the quantity and surface area of placental villi and vessels differ between severe, early-onset fetal growth restriction with absent or reversed umbilical artery Doppler indices and hypertensive disorders of pregnancy or the 2 conditions combined among subjects with disease severity that warrant early preterm delivery. We hypothesized that the trajectories of placental morphogenesis diverge after a common initiating insult of deep defective placentation. Specifically, we postulated that only villi are affected in pregnancy-related hypertension, whereas both villous and vascular structures are proportionally diminished in severe fetal growth restriction with no additional effect when hypertension is concomitantly present. STUDY DESIGN: In this retrospective cohort study, paraffin-embedded placental tissue was obtained from 4 groups, namely (1) patients with severe fetal growth restriction with absent or reversed umbilical artery end-diastolic velocities and hypertensive disorders of pregnancy, (2) patients with severe fetal growth restriction with absent or reversed umbilical artery Doppler indices and no hypertension, (3) gestational age-matched, appropriately grown pregnancies with hypertensive disease, and (4) gestational age-matched, appropriately grown pregnancies without hypertension. Dual immunohistochemistry for cytokeratin-7 (trophoblast) and CD34 (endothelial cells) was performed, followed by artificial intelligence-driven morphometric analyses. The number of villi, total villous area, number of fetoplacental vessels, and total vascular area across villi within a uniform region of interest were quantified. Quantitative analyses of placental structures were modeled using linear regression. RESULTS: Placentas from pregnancies complicated by hypertensive disorders of pregnancy exhibited significantly fewer stem villi (-282 stem villi; 95% confidence interval, -467 to -98; P<.01), a smaller stem villous area (-4.3 mm2; 95% confidence interval, -7.3 to -1.2; P<.01), and fewer stem villous vessels (-4967 stem villous vessels; 95% confidence interval, -8501 to -1433; P<.01) with no difference in the total vascular area. In contrast, placental abnormalities in cases with severe growth restriction were limited to terminal villi with global decreases in the number of villi (-873 terminal villi; 95% confidence interval, -1501 to -246; P<.01), the villous area (-1.5 mm2; 95% confidence interval, -2.7 to -0.4; P<.01), the number of blood vessels (-5165 terminal villous vessels; 95% confidence interval, -8201 to -2128; P<.01), and the vascular area (-0.6 mm2; 95% confidence interval, -1.1 to -0.1; P=.02). The combination of hypertension and growth restriction had no additional effect beyond the individual impact of each state. CONCLUSION: Pregnancies complicated by hypertensive disorders of pregnancy exhibited defects in the stem villi only, whereas placental abnormalities in severely growth restricted pregnancies with absent or reversed umbilical artery end-diastolic velocities were limited to the terminal villi. There were no significant statistical interactions in the combination of growth restriction and hypertension, suggesting that distinct pathophysiological pathways downstream of the initial insult of defective placentation are involved in each entity and do not synergize to lead to more severe pathologic consequences. Delineating mechanisms that underly the divergence in placental development after a common inciting event of defective deep placentation may shed light on new targets for prevention or treatment.

Cyfra 21.1: A Useful Tumour Marker in Pancreatic Ductal Adenocarcinoma: Cross-Sectional Study.

BACKGROUND: Pancreatic cancer is a challenging disease, often requiring invasive procedures for diagnosis. Reliable tumour markers are essential for ensuring early detection and better patient outcomes. Although Carbohydrate Antigen 19-9 is the most commonly used marker, it is marred by low predictive accuracy and high false positivity. Carcino Embryonic Antigen also has limited practical use. A novel antigen, Cytokeratin fragment 21-1, is gaining significance for its diagnostic value in various tumours. MATERIALS AND METHODS: This prospective study aimed to evaluate the potential of Cytokeratin fragment 21-1 in comparison with Carbohydrate Antigen 19-9 and Carcino Embryonic Antigen in diagnosing pancreatic cancer. From January 2016 to December 2019, 45 patients with confirmed pancreatic ductal adenocarcinoma were included in this cross-sectional study. RESULTS: Carbohydrate Antigen 19-9 was raised in 22 patients, Carcino Embryonic Antigen was elevated in 17, and Cytokeratin fragment 21-1 was elevated in 30 cases. Carbohydrate Antigen 19-9 was found to be elevated in the presence of jaundice. Both Carbohydrate Antigen 19-9 and Cytokeratin fragment 21-1 had good correlation with stage of cancer, while Carcino Embryonic Antigen had very minimal correlation. CONCLUSION: In this study, Cytokeratin fragment 21-1 was elevated in a higher number of cases than Carbohydrate Antigen 19-9 and Carcino Embryonic Antigen. Both Cytokeratin fragment 21-1 and Carbohydrate Antigen 19-9 correlated well with cancer stage. Also Cytokeratin fragment 21-1 was not affected by jaundice, unlike Carbohydrate Antigen 19-9. Therefore, Cytokeratin fragment 21-1 has the potential to be an effective individual tumour marker in pancreatic cancer.

Quantification of the lung cancer tumor marker CYFRA 21-1 using protein precipitation, immunoaffinity bottom-up LC-MS/MS.

OBJECTIVES: Numerous studies have proven the potential of cytokeratin 19 fragment 21-1 (CYFRA 21-1) detection in the (early) diagnosis and treatment monitoring of non-small cell lung cancer (NSCLC). Conventional immunoassays for CYFRA 21-1 quantification are however prone to interferences and lack diagnostic sensitivity and standardization. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) is an emerging approach based on a different, often superior, detection principle, which may improve the clinical applicability of CYFRA 21-1 in cancer diagnostics. Therefore, we developed and validated a protein precipitation, immunoaffinity (IA) LC-MS/MS assay for quantitative analysis of serum CYFRA 21-1. METHODS: Selective sample preparation was performed using ammonium sulfate (AS) precipitation, IA purification, tryptic digestion and LC-MS/MS quantification using a signature peptide and isotopically labeled internal standard. The workflow was optimized and validated according to EMA guidelines and results were compared to a conventional immunoassay. RESULTS: Significant interference effects were seen during IA purification, which were sufficiently solved by performing AS precipitation prior to IA purification. A linear calibration curve was obtained in the range of 1.0-100 ng/mL (R2=0.98). Accuracy and precision were well within acceptance criteria. In sera of patients suspected of lung cancer, the method showed good correlation with the immunoassay. CONCLUSIONS: A robust AS precipitation-IA LC-MS/MS assay for the quantification of serum CYFRA 21-1 was developed. With this assay, the clinically added value of LC-MS/MS-based detection over immunoassays can be further explored.

Cytokeratin expression by sarcomas does not indicate synovial origin in dogs.

Sarcoma samples from 33 dogs, 25 subcutaneous and 8 articular, were submitted for cytokeratin immunohistochemistry. Eight of the 25 subcutaneous sarcomas (32%) expressed cytokeratin in 1% to 50% of the neoplastic cells. Of the 7 articular sarcomas evaluated, 1 (14%) expressed cytokeratin in 10% of neoplastic cells. The Kaplan-Meier survival analysis showed that the mean overall survival of dogs with subcutaneous sarcomas (28.1 months [confidence interval [CI]:17.8, 38.4]) did not significantly differ from those with articular sarcomas (24.8 months [CI = 0.5, 29.0]). Overall survival of dogs with sarcomas (both locations combined) immunoreactive for cytokeratin (31.2 months [CI = 17.8, 44.6]) did not differ from those not immunoreactive for cytokeratin (22.0 months [CI = 8.4, 35.6]). Therefore, cytokeratin expression does not indicate synovial origin (P = .64) and neither sarcoma location (P = .76) nor cytokeratin expression (P = .53) affects patient overall survival in this small study. The use of cytokeratin immunohistochemistry is not helpful to determine synovial origin of sarcomas in dogs.

Cytokeratin AE1/AE3 immunolabeling in epithelioid hemangiosarcoma.

Epithelioid hemangiosarcoma (EH), a rare histological variant of hemangiosarcoma, is reported in various animal species, including humans, dogs, cows, horses, and cats. Epithelioid hemangiosarcomas are composed of highly pleomorphic epithelioid cells arranged in cords, islands, nests, or solid cellular areas, similar to epithelial neoplasms. Moreover, in humans, approximately 50% of EHs have cytoplasmic immunolabeling for cytokeratin AE1/AE3 (CK AE1/AE3), making it challenging to distinguish them from carcinomas. This retrospective study assessed the CK AE1/AE3 immunolabeling in canine EH cases from 5 veterinary institutions. Immunohistochemistry for CD31 and CK AE1/AE3 was performed on 30 cases. CK AE1/AE3 immunolabeling was detected in 43% (13/30) of cases, with cytoplasmic labeling ranging from 5% to 100% of neoplastic cells. All tumors consistently had membranous immunolabeling for CD31. The CK AE1/AE3 immunolabeling pattern in canine EHs closely resembled those documented in humans, indicating a similar diagnostic challenge. Therefore, it is recommended to include a vascular immunohistochemistry marker, such as CD31, whenever EH is suspected, particularly in small incisional cutaneous and subcutaneous biopsies.

Cytokeratin-18 is Elevated Prior to Conventional Measures of Liver Disease in Fontan-Associated Liver Disease.

The Fontan procedure is used to palliate complex forms of congenital heart disease. This results in adverse hepatic sequelae now known as Fontan-associated liver disease (FALD). Conventional laboratory measures of liver disease do not correlate well with FALD severity. Cytokeratin-18 (CK-18) is a measure of cell death and is sensitive in detecting other causes of liver disease. Our aim was to assess the use of a novel measure of liver disease, CK-18, in Fontan patients. This is a single-center, prospective, cross-sectional study of Fontan patients aged 8-21 years old. We performed ultrasound elastography, echocardiography, magnetic resonance imaging, and serum laboratory testing. Novel laboratory test CK-18 levels in Fontan subjects were compared to healthy age-matched controls. Thirteen Fontan patients were evaluated with a median age 15 years (10, 14), 4 Hypoplastic left heart syndrome, 11 were male, and 5 were symptomatic. Fontan patients had normal AST/ALT, but a significantly elevated liver stiffness by elastography (median 13.4 kPa). Hepatic stiffness by elastography was associated with diastolic-indexed (rho = 0.58, p = 0.04) ventricular volumes. Compared to 10 aged-matched controls, CK-18 was higher in the Fontan group-cleaved CK-18 protein (p < 0.01) and full CK-18 protein, (p = 0.02). CK-18 was positively associated with AST and ALT. Elevated CK-18 levels were found in Fontan patients compared to controls suggesting hepatic cell death even in these relatively healthy Fontan patients. CK-18 was elevated prior to changes in traditional testing. CK-18 may be a useful sensitive marker of liver disease in FALD.

Unraveling the Keratin Expression in Oral Leukoplakia: A Scoping Review.

Intermediate filaments are one of three polymeric structures that form the cytoskeleton of epithelial cells. In the epithelium, these filaments are made up of a variety of keratin proteins. Intermediate filaments complete a wide range of functions in keratinocytes, including maintaining cell structure, cell growth, cell proliferation, cell migration, and more. Given that these functions are intimately associated with the carcinogenic process, and that hyperkeratinization is a quintessential feature of oral leukoplakias, the utility of keratins in oral leukoplakia is yet to be fully explored. This scoping review aims to outline the current knowledge founded on original studies on human tissues regarding the expression and utility of keratins as diagnostic, prognostic, and predictive biomarkers in oral leukoplakias. After using a search strategy developed for several scientific databases, namely, PubMed, Scopus, Web of Science, and OVID, 42 papers met the inclusion and exclusion criteria. One more article was added when it was identified through manually searching the list of references. The included papers were published between 1989 and 2024. Keratins 1-20 were investigated in the 43 included studies, and their expression was assessed in oral leukoplakia and dysplasia cases. Only five studies investigated the prognostic role of keratins in relation to malignant transformation. No studies evaluated keratins as a diagnostic adjunct or predictive tool. Evidence supports the idea that dysplasia disrupts the terminal differentiation pathway of primary keratins. Gain of keratin 17 expression and loss of keratin 13 were significantly observed in differentiated epithelial dysplasia. Also, the keratin 19 extension into suprabasal cells has been associated with the evolving features of dysplasia. The loss of keratin1/keratin 10 has been significantly associated with high-grade dysplasia. The prognostic value of cytokeratins has shown conflicting results, and further studies are required to ascertain their role in predicting the malignant transformation of oral leukoplakia.

Blastocyst-like Structures in the Peripheral Retina of Young Adult Beagles.

In this immunohistological study on the peripheral retina of 3-year-old beagle dogs, excised retina specimens were immunostained with antibodies against nestin, Oct4, Nanog, Sox2, CDX2, cytokeratin 18 (CK 18), RPE65, and YAP1, as well as hematoxylin and DAPI, two nuclear stains. Our findings revealed solitary cysts of various sizes in the inner retina. Intriguingly, a mass of small round cells with scant cytoplasms was observed in the cavity of small cysts, while many disorganized cells partially occupied the cavity of the large cysts. The small cysts were strongly positive for nestin, Oct4, Nanog, Sox2, CDX2, CK18, and YAP1. RPE65-positive cells were exclusively observed in the tissue surrounding the cysts. Since RPE65 is a specific marker of retinal pigment epithelial (RPE) cells, the surrounding cells of the peripheral cysts were presumably derived from RPE cells that migrated intraretinally. In the small cysts, intense positive staining for nestin, a marker of retinal stem cells, seemed to indicate that they were derived from retinal stem cells. The morphology and positive staining for markers of blastocyst and RPE cells indicated that the small cysts may have formed structures resembling the blastocyst, possibly caused by the interaction between retinal stem cells and migrated RPE cells.

Gastric Metastasis Mimicking Early Gastric Cancer from Invasive Ductal Carcinoma of the Breast: Case Report and Literature Review.

Backgound and Objectives: Gastric metastasis from invasive ductal breast cancer (BC) is rare. It mainly occurs in patients with lobular BC. The occurrence of multiple metastases is typically observed several years after the primary diagnosis. Endoscopic findings of gastric metastasis of the BC were usually the linitis plastic type. Case presentation: A 72-year-old women who underwent right modified radical mastectomy (MRM) 10 month ago was referred after being diagnosed with early gastric cancer (EGC) during systemic chemotherapy. EGC type I was found at gastric fundus, and pathologic finding showed poorly differentiated adenocarcinoma. Metachronous double primary tumor EGC was considered. Management and Outcome: A laparoscopic total gastrectomy was performed, and postoperative pathology revealed submucosa invasion and two lymph node metastases. A pathologic review that focused on immunohistochemical studies of selected antibodies such as GATA binding protein 3 (GATA3), gross cystic disease fluid protein-15 (GCDFP-15), cytokeratin 7 (CK7) was performed again, comparing previous results. As a result, gastric metastasis from BC was diagnosed. After totally laparoscopic total gastrectomy, palliative first-line chemotherapy with paclitaxel/CDDP was performed. Two months after gastrectomy, she was diagnosed with para-aortic lymph node metastasis and multiple bone metastases. She expired six months after gastrectomy. Conclusions: Gastric metastasis from invasive ductal carcinoma of the breast, which is clinically manifested as EGC, is a very rare condition. If there is a history of BC, careful pathological review will be required.

Expression and diagnostic value of serum free light chain in lung cancer. / è¡€æ¸ æ¸¸ç¦»è½»é“¾åœ¨è‚ºç™Œä¸­çš„è¡¨è¾¾åŠè¯Šæ–­ä»·å€¼.

OBJECTIVES: The expression of serum free light chain (FLC) is abnormal in various diseases, but its role in lung cancer remains unclear. This study aims to investigate the expression and diagnostic value of serum FLC in lung cancer. METHODS: A total of 80 lung cancer patients treated at Xiangdong Hospital, Hunan Normal University from January to December 2021 were selected as the lung cancer group. Another 80 healthy individuals undergoing routine physical examinations during the same period were chosen as the control group. General information and serum κFLC and λFLC levels were collected for all subjects. Clinical indicators such as serum carcinoembryonic antigen (CEA), cytokeratin fragment antigen 21-1 (CYFRA21-1) levels, tumor diameter, histological type, TNM stage, and lymph node metastasis status were recorded for lung cancer patients. The expression levels of serum FLC [κFLC, λFLC, and FLC (κ+λ)] were compared between the lung cancer group and the control group. Lung cancer patients were grouped based on gender, age, smoking history, tumor diameter, TNM stage, histological type, and lymph node metastasis to compare differences in serum κFLC and λFLC levels. Receiver operating characteristic (ROC) curves were used to evaluate the diagnostic value of serum FLC alone and in combination with other indicators in lung cancer. RESULTS: The expression levels of serum FLC (κ+λ) and κFLC were significantly higher in the lung cancer group than those in the control group (both P<0.001), while there was no significant difference in serum λFLC levels between the 2 groups (P>0.05). There were no significant differences in serum κFLC levels among lung cancer patients with different tumor diameters, histological types, or TNM stages (all P>0.05); however, serum κFLC levels were higher in lung cancer patients with lymph node metastasis than in those without, with statistical significance (P=0.033). There were no significant differences in serum λFLC levels based on tumor diameter or histological type (both P>0.05), but serum λFLC levels were higher in stage III+IV and lymph node metastatic lung cancer patients compared to stage I+II and non-metastatic patients, with statistical significance (P=0.033 and P=0.019, respectively). The area under the curve (AUC) for κFLC and CEA in diagnosing lung cancer showed no significant difference (P=0.333). The combination of κFLC+CYFRA21-1 had the highest diagnostic efficacy (AUC=0.875) and sensitivity (71.3%). The AUC for the combined diagnosis of κFLC+λFLC+CEA+CYFRA21-1 was 0.915 (95% CI 0.860 to 0.953, P<0.001). CONCLUSIONS: Serum FLC is highly expressed in lung cancer and is associated with its invasion and metastasis. Serum FLC, particularly κFLC, has diagnostic value for lung cancer, and the combined detection of FLC, CEA, and CYFRA21-1 offers the best diagnostic efficacy.

Vimentin and cytokeratin: Good alone, bad together.

The cytoskeleton plays an integral role in maintaining the integrity of epithelial cells. Epithelial cells primarily employ cytokeratin in their cytoskeleton, whereas mesenchymal cells use vimentin. During the epithelial-mesenchymal transition (EMT), cytokeratin-positive epithelial cells begin to express vimentin. EMT induces stem cell properties and drives metastasis, chemoresistance, and tumor relapse. Most studies of the functions of cytokeratin and vimentin have relied on the use of either epithelial or mesenchymal cell types. However, it is important to understand how these two cytoskeleton intermediate filaments function when co-expressed in cells undergoing EMT. Here, we discuss the individual and shared functions of cytokeratin and vimentin that coalesce during EMT and how alterations in intermediate filament expression influence carcinoma progression.

Expression of high molecular weight cytokeratin-A novel feature of aggressive and innate hormone-refractory prostatic adenocarcinoma.

BACKGROUND: Castration-resistance is common in advanced prostatic adenocarcinomas (PACs) treated with androgen deprivation therapy (ADT) and usually occurs after 2 years following treatment. A minority of PACs confer innate ADT resistance without prior hormonal treatment. The expression of HMWCK in PAC cells has not been studied. This study aimed to investigate the clinicopathologic and genomic features of HMWCK-expressing PACs and the relationship to ADT resistance. METHODS: A total of 469 PACs were studied for HMWCK expression (39 postradiotherapy, 57 post-ADT, 373 treatment-naïve PACs). Clinicopathologic correlations of the HMWCK expression with tumor grade groups, specific tumor morphologies, tumor stages and disease recurrence/persistence/progression were performed. Five HMWCK+ PACs were also sequenced for genetic alterations. RESULTS: Thirty one of the 469 cases (6.6%) showed variable HMWCK+ PAC. The HMWCK+ PAC often focally presented in the tumor and vast majority were associated with high Gleason scores and unfavorable growth patterns (cribriform, comedo-necrosis, and intraductal carcinoma) as well as high tumor stages. A small percentage of the HMWCK+ PCA (2/31, 6.5%) presented with frank squamous histomorphology. Vast majority (22/31, 87%) had no history of prior ADT. The HMWCK+ PAC all displayed diminished to lost expression of AR/NKX3.1. Most of the cases progressed within 12 months of ADT or disease persisted despite ADT. Of the 5 HMWCK+ PACs subjected to gene sequencing, 4 presented with PTEN/PI3K/MAPK pathway alterations. CONCLUSION: The study demonstrated HMWCK+ PAC to be a novel type of innate ADT-resistant PAC. Overexpression of HMWCK in PAC can be potentially used as a surrogate biomarker for aggressive and innate hormone-refractory PACs. The genetic alterations imply potential therapeutic implications of PI3K/MAPK inhibitors in the treatment of these deadly diseases.

Keratin 79 is a PPARA target that is highly expressed by liver damage.

Keratins are key structural proteins found in skin and other epithelial tissues. Keratins also protect epithelial cells from damage or stress. Fifty-four human keratins were identified and classified into two families, type I and type II. Accumulating studies showed that keratin expression is highly tissue-specific and used as a diagnostic marker for human diseases. Notably, keratin 79 (KRT79) is type II cytokeratin that was identified as regulator of hair canal morphogenesis and regeneration in skin, but its role in liver remains unclear. KRT79 is undetectable in normal mouse but its expression is significantly increased by the PPARA agonist WY-14643 and fenofibrate, and completely abolished in Ppara-null mice. The Krt79 gene has functional PPARA binding element between exon 1 and exon 2. Hepatic Krt79 is regulated by HNF4A and HER2. Moreover, hepatic KRT79 is also significantly elevated by fasting- and high-fat diet-induced stress, and these increases are completely abolished in Ppara-null mice. These findings suggest that hepatic KRT79 is controlled by PPARA and is highly associated with liver damage. Thus, KRT79 may be considered as a diagnostic marker for human liver diseases.

Antibody against apolipoprotein-A1, non-alcoholic fatty liver disease and cardiovascular risk: a translational study.

BACKGROUND: Non-alcoholic fatty liver disease (NAFLD) is a common liver disease increasing cardiovascular disease (CVD) morbidity and mortality. Autoantibodies against apolipoprotein A-1 (AAA-1) are a possible novel CVD risk factor promoting inflammation and disrupting cellular lipid homeostasis, two prominent pathogenic features of NAFLD. We explored the role of AAA-1 in NAFLD and their association with CVD risk. METHODS: HepaRG cells and liver sections from ApoE-/- mice exposed to AAA-1 were used for lipid quantification and conditional protein expression. Randomly selected sera from 312 subjects of the Prevention of Renal and Vascular End-stage Disease (PREVEND) general population cohort were used to measure AAA-1. A Fatty Liver Index (FLI) ≥ 60 and a 10-year Framingham Risk Score (FRS) ≥ 20% were used as proxy of NAFLD and high CVD risk, respectively. RESULTS: In-vitro and mouse models showed that AAA-1 increased triglyceride synthesis leading to steatosis, and promoted inflammation and hepatocyte injury. In the 112 PREVEND participants with FLI ≥ 60, AAA-1 were associated with higher FRS, alkaline phosphatase levels, lower HDL cholesterol and tended to display higher FLI values. Univariate linear and logistic regression analyses (LRA) confirmed significant associations between AAA-1, FLI and FRS ≥ 20%, while in adjusted LRA, FLI was the sole independent predictor of FRS ≥ 20% (OR: 1.05, 95%CI 1.01-1.09, P = 0.003). AAA-1 was not an independent FLI predictor. CONCLUSIONS: AAA-1 induce a NAFLD-compatible phenotype in vitro and in mice. Intricate associations exist between AAA-1, CVD risk and FLI in the general population. Further work is required to refine the role of AAA-1 in NAFLD and to determine if the AAA-1 association with CVD is affected by hepatic steatosis.

The increased expression of cytokeratin 13 leads to an increase in radiosensitivity of nasopharyngeal carcinoma HNE-3 cells by upregulating ERRFI1.

The main factors contributing to the unfavorable outcome in the clinical treatment of patients with nasopharyngeal carcinoma (NPC) patients are radiation resistance and recurrence. This study aimed to investigate the sensitivity and molecular foundation of cytokeratin 13 (CK13) in the radiotherapy of NPC. To achieve this, a human NPC cell line overexpressing CK13, HNE-3-CK13, was constructed. The effects of CK13 overexpression on cell viability and apoptosis under radiotherapy conditions were evaluated using the CCK-8 assay, immunofluorescence, and western blotting (WB). Next-generation sequencing was performed to identify the downstream genes and signaling pathways of CK13 that mediate radiotherapy response. The potential role of the candidate gene ERRFI1 in CK13-induced enhancement of radiosensitivity was investigated through rescue experiments using clone formation and WB. The effects of ERRFI1 on cell viability, cell apoptosis, cell cycle, and the related key genes were further evaluated using CCK-8, immunofluorescence, flow cytometry, quantitative polymerase chain reaction and WB. The results showed that CK13 overexpression in HNE-3 significantly inhibited cell survival under radiotherapy and promoted apoptosis marker γH2AX expression, leading to a significant increase of ERRFI1. Knockdown of ERRFI1 rescued the decreased cell viability and proliferation and the increased cell apoptosis that were caused by CK13 overexpression-mediated radiotherapy sensitization of NPC cells. In this process, EGFR, AKT, and GSK-3ß were found involved. In the end, ERRFI1 was proven to inhibit expression levels of CDK1, CDK2, cyclin B1, and cyclin D1, resulting an increased G2/M cell ratio. Overexpression of CK13 enhances the radiosensitivity of NPC cells, which is characterized by decreased cell viability and proliferation and increased apoptosis. This regulation may affect the survival of HNE-3 cells by increasing the expression of ERRFI1 and activating the EGFR/Akt/GSK-3ß signaling pathway, providing new potential therapeutic targets for the treatment of NPC.