Loss of protein kinase calpha expression may enhance the tumorigenic potential of Gli1 in basal cell carcinoma

Gli1 has now been implicated in switching between motility and static cell replication. Part of a series of papers, this study tracted PKCalpha expression in relation to Gli1 in various static and motile cell areas of the hair sheath and BCC

Exploiting the efficacy of Tyro3 and folate receptors to enhance the delivery of gold nanoparticles into colorectal cancer cells in vitro

Colorectal cancer (CRC) is the fourth most common cancer in the world. Due to its asymptomatic nature, CRC is diagnosed at an advanced stage where the survival rate is <5%. Besides, CRC treatment using chemotherapy, radiotherapy and surgery often causes undesirable side-effects. As such, gold nanoparticles (GNPs) are envisaged in the field for the diagnosis and treatment of CRC. GNPs have unique physical, chemical and electrical properties at the nanoscale which make them suitable for application in biomedicine. However, for GNPs to become clinically effective, their internalisation efficiency in cancer cells must be enhanced. Folate receptor-α (FR) is overexpressed in CRC cells wherein FR helps in the uptake of folic acid within the cells. Tyro3, a novel tyrosine kinase receptor, drives cell proliferation and its overexpression is correlated with poor prognosis in CRC. Their upregulated expression in CRC cells relative to normal cells makes them an ideal target for GNPs using active targeting. Therefore, in this study receptors FR and Tyro3 were simultaneously targeted using specific antibody-coated GNPs in order to enhance the uptake and internalisation of GNPs in CRC cells in vitro. Four different types of coated-GNPs were synthesised GNPs-PEG, GNPs-anti-FR, GNPs-anti-Tyro3 and GNPs-anti-(FR + Tyro3) and incubated (0–50 ng) with three CRC cell lines namely CRL1790, CRL2159 and HCT116. Simultaneous targeting of these receptors by GNPs-anti-(FR + Tyro3) was found to be the most effective in internalisation in CRC cells compared with GNPs targeted singly to FR or Tyro3 (p <0.05). Besides this, results show that Tyro3 mediated similar internalisation efficacy to FR (p <0.05) in CRC cells using ICP-OES

Recent advances in arsenic trioxide encapsulated nanoparticlesas drug delivery agents to solid cancers

Since arsenic trioxide (ATO) was first approved as the front line therapy for acute promyelocytic leukemia (APL) 25 years ago, its anti-cancer properties for various malignancies have been under intense investigation. However, the clinical successes of ATO in treating hematological cancers have not been translated to solid cancers. This is due to arsenic’s rapid clearance by the body’s immune system before reaching the tumor site. Several attempts have henceforth been made to increase its bioavailability toward solid cancers without increasing its dosage albeit without much success. This review summarizes the past and current utilization of ATO in the medical field with primary focus on the implementation of nanotechnology for ATO delivery to solid cancer cells. Different approaches that have been employed to increase arsenic’s efficacy, specificity and bioavailability to solid cancer cells were evaluated and compared. The potential of combining different approaches or tailoring delivery vehicles to target specific types of solid cancers according to individual cancer characteristics and arsenic chemistry was proposed and discussed

Modelling and validating three dimensional human normal cervix and cervical cancer tissues in vitro

Objective: The use of three dimensional in vitro systems in cancer research is a promising path for developing effective anticancer therapies. The aim of this study was to engineer a functional 3-D in vitro model of normal and cancerous cervical tissue. Methods: Normal epithelial and immortalized cervical epithelial carcinoma cell lines were used to construct 3-D artificial normal cervical and cervical cancerous tissues. De-epidermised dermis (DED) was used as a scaffold for both models. Morphological analyses were conducted by using haematoxylin and eosin staining and characteristics of the models were studied by analysing the expression of different structural cytokeratins and differential protein marker Mad1 using immunohistochemical technique. Results: Haematoxylin and eosin staining results showed that normal cervical tissue had multi epithelial layers while cancerous cervical tissue showed dysplastic changes. Immunohistochemistry staining results revealed that for normal cervix model cytokeratin 10 was expressed in the upper stratified layer of epithelium while cytokeratin 5 was expressed mainly in the middle and basal layer. Cytokeratin 19 was weakly expressed in a few basal cells. Cervical cancer model showed cytokeratin 19 expression in different epithelial layers and weak or no expression for cytokeratin 5 and cytokeratin 10. Mad1 expression was detected in some suprabasal cells. Conclusions: The 3-D in vitro models showed stratified epithelial layers and expressed the same types and patterns of differentiation marker proteins as seen in corresponding in vivo tissue in either normal cervical or cervical cancerous tissue. Findings imply that they can serve as functional normal and cervical cancer models

The role of ectopic human chorionic gonadotropin beta subunit in inducing epithelial mesenchymal transition in human keratinocytes and its possible pathways

Background: The process of epithelial-mesenchymal transition (EMT) involves the trans-differentiation of epithelial cells to mesenchymal cells associated with high plasticity. It usually occurs when the cells acquire migratory and invasive characteristics due to the weakening or the loss of cell-cell adhesion. Human chorionic gonadotropin (hCG), a pregnancy hormone, consists of a common α subunit which is shared by three other hormones, thyroid stimulating hormone, luteinizing hormone and follicular stimulating hormone; and an unique β subunit (hCGβ). Previous studies have demonstrated that hCGβ was expressed by some epithelial origin cancers (1, 2, 3) and therefore it has been postulated as a possible epithelial cancer biomarker. Other studies have linked the presence hCGβ to the aggressive and invasive behavior of certain cancers and their poor prognosis (3, 4). Methods: This study was set out to investigate whether hCGβ plays a role in inducing the EMT and to elucidate the possible pathways. Human keratinocytes (HK) were exposed to spent media collected from hCGβ producing cancer cells (ScaBER cells) for 48 hours before the cells were either fixed for immnuostaining or cells were lysed and protein extracts were collected for western blotting analysis. The expression of epithelial and mesenchymal markers was evaluated by both florescent immunocytochemistry and western blotting techniques. Results: A trend of up-regulation of mesenchymal markers (Vimentin and β-catenin) and down regulation of epithelial marker (E-cadherin) in these treated HK cells was observed. There was 50% increase in cell number which was positively stained by anti-Vimentin antibody whilst 16% of the cells have lost E-cadherin expression (100% to 84%) following 48 hours’ exposure to the hCGβ containing media. These findings were in consistence with the results from HK cells that were exposed to recombinant hCGβ (r-hCGβ). It was also observed that the changes in the expressions of these markers were reduced when a combination of three anti-hCGβ antibodies targeting different hCGβ epitopes was added to the spent media. These results were confirmed by western blotting analysis. Conclusion: The findings suggest that ectopic hCGβ produced by cancer cells might be involved in EMT associated with the migratory and aggressive behavior of such cancers. Furthermore, the up-regulation of β-catenin also suggests its possible role in the Wnt pathway which offers an insight into EMT process at a molecular level. This could be valuable point in developing future novel anti hCGβ therapies for such types of cancers

Effective delivery of arsenic trioxide to HPV-positive cervical cancer cells using optimised liposomes: a size and charge study

Despite the success of arsenic trioxide (ATO) in treating haematological malignancies, its potential to treat solid tumours has not been fully exploited, owing to its dose-limiting toxicity and poor pharmacokinetics. In order to overcome this hurdle, liposomal encapsulation of the drug with different surface charges (neutral, negative, and positive) and sizes (100, 200 and 400 nm) were synthesised and tested on human papilloma virus (HPV)-positive HeLa and HPV-negative HT-3 cervical cancer cell lines. Two epithelial cell lines-human keratinocytes (HK) and human colon cells (CRL-1790)-were used as controls. The synthesised liposomes were tested for their physico-chemical characteristics, drug loading efficiency, and toxicity on the studied cell lines. Neutral liposomes of 100 nm in size were the chosen formulation for delivering ATO into the studied cells, as they showed the least intrinsic cytotoxicity and the highest loading efficiency. The findings demonstrated that the optimised formulation of liposomes was an effective drug delivery method for HPV-infected cervical cancer cells. Furthermore, the toxicity vs. uptake ratio was highest for HeLa cells, while a reduced or minimal toxic effect was observed for non-HPV-infected cervical cancer cells and control cells. These findings may provide a promising therapeutic strategy for effectively managing cervical cancers

Positive correlational shift between crevicular antimicrobial peptide LL-37, pain and periodontal status following non-surgical periodontal therapy. A pilot study

Background: Periodontitis has a high prevalence and uncertain recurrence. Unlike the pro-inflammatory cytokine profile, little is known about the anti-inflammatory cytokine and antimicrobial peptide overview following treatment. The present study aimed to evaluate if any of the antimicrobial peptide LL-37, interleukin (IL) 4, 10 and 6 together with the volume of gingival crevicular fluid (GCF) and total protein concentration in GCF could be used as correlative biomarkers for the severity in periodontitis as well as prognostic factors in the management of the disease. Methods: Forty-five participants were recruited and allocated to the healthy (15), Stage I-II (15) or Stage III-IV periodontitis (15) group. Along with periodontal examination, GCF samples were obtained at baseline and 4–6 weeks following scaling and root planing (SRP) for the periodontitis groups. GCF samples were analyzed by ELISA kits to quantify LL-37 and IL-4, -6 and − 10. One-way ANOVA followed by Dunnett’s test was used to determine differences among the three groups at baseline. Two-way ANOVA followed by Sidak’s post-hoc test was used to compare between pre- and post-SRP in the two periodontitis groups. Results: The amount of GCF volume was significantly correlated to the severity of periodontitis and decreased following SRP, particularly in the Stage III-IV group (p < 0.01). The levels of LL-37, IL-6, and pain and periodontal clinical parameters were significantly correlated to the severity of periodontitis. IL-4 and IL-10 in the periodontitis groups were significantly lower than the healthy group (p < 0.0001) and barely improved following SRP up to the level of the healthy group. Conclusions: With the limitations of this study, crevicular LL-37 may be a candidate for a biomarker of periodontitis and the associated pain upon probing. Trial registration: The study was registered in clinical trials.gov, with number NCT04404335, dated 27/05/2020

Optimisation of folate-mediated liposomal encapsulated arsenic trioxide for treating HPV-positive cervical cancer cells in vitro

High-risk human papilloma virus (HPV) infection is directly associated with cervical cancer development. Arsenic trioxide (ATO), despite inducing apoptosis in HPV-infected cervical cancer cells in vitro, has been compromised by toxicity and poor pharmacokinetics in clinical trials. Therefore, to improve ATO’s therapeutic profile for HPV-related cancers, this study aims to explore the effects of length of ligand spacers of folate-targeted liposomes on the efficiency of ATO delivery to HPV-infected cells. Fluorescent ATO encapsulated liposomes with folic acid (FA) conjugated to two different PEG lengths (2000 Da and 5000 Da) were synthesised, and their cellular uptake was examined for HPV-positive HeLa and KB and HPV-negative HT-3 cells using confocal microscopy, flow cytometry, and spectrophotometer readings. Cellular arsenic quantification and anti-tumour efficacy was evaluated through inductively coupled plasma-mass spectrometry (ICP-MS) and cytotoxicity studies, respectively. Results showed that liposomes with a longer folic acid-polyethylene glycol (FA-PEG) spacer (5000 Da) displayed a higher efficiency in targeting folate receptor (FR) + HPV-infected cells without increasing any inherent cytotoxicity. Targeted liposomally delivered ATO also displayed superior selectivity and efficiency in inducing higher cell apoptosis in HPV-positive cells per unit of arsenic taken up than free ATO, in contrast to HT-3. These findings may hold promise in improving the management of HPV-associated cancers

Modelling and validating three-dimensional human breast and cancerous human breast tissues in vitro

In this study three dimensional (3-D) in vitro models of normal breast and breast cancer tissues were developed to mimic closely the in vivo tissue microenvironment and therefore providing reliable models for in vitro studies as well as testing of novel cancer therapies. Normal and cancerous human breast cell lines were used to construct 3-D artificial tissues, where de-epidermalised dermis (DED) was used as a scaffold for both models. Morphological analyses were conducted using haematoxylin and eosin staining. Biomarkers including keratin 5 and 19 as well as α smooth muscle actin and mucin 1 were used to confirm and validate the reliability of the proposed models using immunohistochemical techniques. Findings suggest that the 3-D in vitro models described in this work can serve as functional models of both human normal and cancerous breast tissues. Multiple structures similar to ducts and lobules of human breast in vivo were observed in 3-D in vitro models by the use of H&E, some breast cancer colonies seen in the cancerous 3-D model were similar to the ducto-lobular structures observed in normal 3-D model of the breast but the former cells were more loosely connected, irregular and largely disorganized. The established 3-D in vitro model of normal breast showed the development of ducto-lobular structures composed of an inner cell layer which was stained positive with α mucin 1 antibody, a biomarker that is characteristic for luminal cells; and also an outer basal layer of cells that was stained positive for α smooth muscle actin, a biomarker of myoepithelial cells.. Keratin staining in 3-D in vitro models also resembled the pattern observed in vivo where keratin 5 was detected in both luminal and myoepithelial cells of normal breast model (NTERT cells), whereas keratin 19 was present in breast cancer model (C2321 cells). These 3-D models successfully recapitulate both normal and pathological tissue architecture of breast tissue and has the potential for various applications in the evaluation of breast cancer progression and treatment