Therapeutic Prospects in Preeclampsia - A Mini-Review.

BACKGROUND: Preeclapmsia (PE) is characterized by early onset symptoms such as elevated blood pressure, proteinuria and edema in the pregnant woman, and may result in seizures in the affected female. Currently, there are no therapeutic drugs available to treat this condition, but there are interventions to regulate the symptoms based on the gestational period of the fetus, although the largely favored option is delivery of the fetus and placenta. OBJECTIVE: A search for biomolecules associated with PE was conducted so as to identify diagnostic markers and therapeutic leads. RESULTS: The literature search resulted in the identification of biomolecules such as Corin and Placental Protein 13 (PP13), among others that are associated with PE. Thereby, giving an insight into the various mechanistic pathways involved in the causation of PE. However, it is also evident that PE cannot be solely attributed to any single mechanism but is due to an interplay of different factors that have led to the development of this disease condition. CONCLUSION: The identified biomarkers would ultimately help in understanding this complex disease and perhaps lead to the discovery of potential effective molecular targets for clinical trials, thereby providing a valuable therapeutic option for affected pregnant women.

Silver nanoparticles in cancer: therapeutic efficacy and toxicity.

In recent years, there has been escalating interest in the biomedical applications of nanoparticles (NPs). In particular, silver nanoparticles (AgNPs) are increasingly being investigated as tools for novel cancer therapeutics, capitalizing on their unique properties to enhance potential therapeutic efficacy. However, questions as to are we able to contain or control the toxicity effects of AgNPs, and how much do we know about the toxicological profile of AgNPs which are commonly used in emerging nanotechnology-based applications, still remain. Hence, serious considerations have to be given to the hazards and risks of toxicity associated with the use of AgNPs. This review focuses on the current applications of AgNPs, their known effects and toxicity, as well as the potential of harnessing them for use in cancer therapy.

Parafibromin expression in breast cancer: a novel marker for prognostication?

BACKGROUND: Parafibromin is a novel protein product of HRPT2, a recently identified tumour suppressor gene. Mutations of the HRPT2 gene are common in parathyroid carcinomas, and these exhibit reduced protein expression. Parafibromin expression in breast cancer has not been previously studied. AIMS: To determine the distribution of parafibromin in breast cancer tissues, and correlate its expression with conventional pathological parameters. METHODS: Tissue microarrays were constructed from archival paraffin embedded breast cancer samples. Sections cut from tissue microarray blocks were subjected to immunohistochemistry. Immunopositivity for parafibromin and intensity-percentage scores were derived by blinded evaluation. Findings were correlated with clinicopathological parameters. RESULTS: 163 breast cancers were assessed. Larger tumours were less likely to express parafibromin than smaller ones, with the association approaching statistical significance (p = 0.05). Staining intensity correlated inversely with tumour size (p = 0.016) and pathological stage (p = 0.008); as did parafibromin intensity-percentage score with pathological stage (p = 0.03), lymphovascular invasion (p = 0.03) and cerbB2 intensity-percentage score (p = 0.04). CONCLUSION: Parafibromin in breast cancer, as in parathyroid tumours, appears to have tumour suppressor functions, with loss of protein expression associated with adverse pathological parameters. These findings may indicate a potential role of parafibromin as a prognostic marker in breast cancer.

Mucinous breast lesions: diagnostic challenges.

Breast lesions with mucin represent a broad spectrum of entities, ranging from benign fibrocystic changes with luminal mucin to mucocele-like lesions (MLL), which can be associated with banal epithelial alterations, atypical ductal hyperplasia or ductal carcinoma in situ. Occasionally invasive mucinous carcinoma can be identified in contiguity with MLL. Diagnostic challenges are enumerated, histological differentials are discussed, and a practical approach towards resolving some of these issues is provided. In addition to these lesions with abundant extracellular mucin, there are also conditions that feature stromal mucinous or myxoid material, as well as rare entities that demonstrate both epithelial extracellular and stromal mucin.

Hypericin photoactivation triggers down-regulation of matrix metalloproteinase-9 expression in well-differentiated human nasopharyngeal cancer cells.

Recently, we have shown that hypericin-mediated photodynamic therapy (PDT) is a promising modality for the treatment of nasopharyngeal cancer (NPC). The present study evaluated the expression of matrix metalloproteinase-9 (MMP-9) following hypericin-PDT in well-differentiated HK1 NPC cells. Down-regulation of MMP-9 by hypericin-PDT was observed at the mRNA level in HK1 cells in vitro and in vivo and at the protein level in vitro. Transcriptional activities of the activator protein-1 (AP-1) and nuclear factor (NF)-kappaB regulatory elements were inhibited by PDT. We also found that PDT reduced secreted granulocyte-macrophage colony stimulating factor (GM-CSF), which is known to activate transcription of NK-kappaB and AP-1. However, incubation of untreated HK1 cells with exogenous GM-CSF abrogated the reduction of MMP-9 production in hypericin-PDT-treated cells. It would appear that PDT downregulates MMP-9 expression via inhibition of GM-CSF production, which in turn modulates AP1/NF-kappaB transcriptional activities. Suppression of MMP-9 by hypericin-PDT may have therapeutic implications.

Evaluation of electrospun PCL/gelatin nanofibrous scaffold for wound healing and layered dermal reconstitution.

The current design requirement for a tissue engineering skin substitute is that of a biodegradable scaffold through which fibroblasts can migrate and populate. This artificial "dermal layer" needs to adhere to and integrate with the wound, which is not always successful for the current artificial dermal analogues available. The high cost of these artificial dermal analogues also makes their application prohibitive both to surgeons and patients. We propose a cost-effective composite consisting of a nanofibrous scaffold directly electrospun onto a polyurethane dressing (Tegaderm, 3M Medical) - which we call the Tegaderm-nanofiber (TG-NF) construct - for dermal wound healing. Cell culture is performed on both sides of the nanofibrous scaffold and tested for fibroblast adhesion and proliferation. It is hoped that these studies will result in a fibroblast-populated three-dimensional dermal analogue that is feasible for layered applications to build up thickness of dermis prior to re-epithelialization. Results obtained in this study suggest that both the TG-NF construct and dual-sided fibroblast-populated nanofiber construct achieved significant cell adhesion, growth and proliferation. This is a successful first step for the nanofiber construct in establishing itself as a suitable three-dimensional scaffold for autogenous fibroblast populations, and providing great potential in the treatment of dermal wounds through layered application.

Nuclear morphometry in columnar cell lesions of the breast: is it useful?

AIMS: To evaluate the nuclear morphometric features of breast columnar cell lesions (CCLs) observed on mammotome core biopsies, to determine if there are significant measurable differences between those with atypia and those without. Correlation with follow-up open excision specimens was made. METHODS: Mammotome core biopsies performed on patients that contained CCLs were derived from the departmental case files. Histological material was reviewed and foci of CCLs demarcated for nuclear morphometric assessment, which was accomplished using an imaging system. Nuclear parameters studied were nuclear area and perimeter, circularity factor and feret's diameter. Statistical analysis used the GraphPad Prism software, with p<0.05 indicating significance. RESULTS: On examination of core biopsies of 40 patients with CCLs, 8 lesions were benign, 4 showed atypical lobular hyperplasia, 8 showed CCLs with nuclear atypia, 19 disclosed atypical ductal hyperplasia (ADH) and 1 showed ductal carcinoma in situ (DCIS). The nuclear area, perimeter and feret's diameter of CCLs with atypia were significantly greater than those without (p = 0.04, 0.03 and 0.019, respectively), whereas no difference was observed in the circularity factor. Follow-up open excision biopsy specimens in 24 patients showed upgrading to DCIS in 40% of cases diagnosed initially with ADH on core biopsy compared with 20% of CCLs with atypia. CONCLUSIONS: Nuclear morphometry in CCLs confirms nuclear size as the key parameter in the assessment of nuclear atypia. Whether it can be potentially used as an adjunctive tool depends on the establishment of appropriate cut-offs.

Up-regulation of metallothionein isoforms in keloid keratinocytes.

Keloids are proliferative growths of dermal collagen, usually resulting from excessive tissue response during wound healing. There is evidence that keratinocytes may promote keloidogenesis via epithelial-mesenchymal interactions. Metallothioneins (MTs) are known to be involved in the fundamental cellular processes of growth and apoptosis. In this study, we evaluated the expression of MT isoforms in normal and keloid keratinocytes. The expression patterns of ten functional MT isoforms were assessed using real-time RT-PCR in primary cultures of normal and keloid keratinocytes. The MT-2A isoform was the most abundant MT isoform in both normal and keloid keratinocytes while the MT-1B isoform was absent. There was a significant increase in the mRNA expression of four MT isoforms, viz. MT-1A, 1E, 1F and 2A in keloid keratinocytes as compared to normal keratinocytes. Up-regulation of MT-1A, 1E, 1F and 2A isoforms may play a part in the development of keloids by paracrine signaling.

Evaluation of cell culture on the polyurethane-based membrane (Tegaderm): implication for tissue engineering of skin.

BACKGROUND: The use of polymer-based delivery systems, on which cells are cultured and transferred, improves the ease of handling and transfer of the keratinocytes. A transparent polymer also allows observation of cell growth prior to grafting as well as re-epithelialization after grafting to the wound. We have developed techniques for cultured keratinocytes on Tegaderm (3M), an inexpensive and easily available polyurethane-based wound dressing, for treatment of burn and chronic wounds. In this study, we evaluate cell culture characteristics of three different cell types, human epidermal keratinocytes, human dermal fibroblasts and pig bone marrow mesenchymal stem cells on Tegaderm membrane. METHODS: Cells were isolated from human skin or pig bone marrow and cultured on membranes for a period of five days. Cell proliferation was assessed by colorimetric assay (MTT) and scanning electron microscopy. RESULTS AND CONCLUSIONS: This study confirms that Tegaderm membranes support attachment and growths for these cell types, with those growth characteristics are similar, if not as good as that of optimal condition of tissue culture plastics. Data from our study suggest that Tegaderm membranes can be used, modified and developed further as an economical and easily available material for tissue engineered skin.

Chondroitin sulfate in palatal wound healing.

Chondroitin sulfate is up-regulated in granulation tissue during wound healing. To investigate the role of chondroitin sulfate in the wound-healing process after surgical repair of cleft palate, we isolated and cultured rabbit palatal fibroblasts. Treatment with chondroitin-6-sulfate resulted in a dose-dependent increase in cell adhesion and cell proliferation, whereas the reverse effects were seen after chondroitinase degradation of chondroitin sulfate. The biological actions of chondroitin sulfate appeared to be dependent on the presence and position of sulfate groups. Inhibition of glycosaminoglycan sulfation by chlorate treatment led to reduced cell adhesion and cell proliferation and a slower rate of wound closure in vitro. Furthermore, exposure to chondroitin-4-sulfate resulted in a dose-dependent reduction in cell adhesion. Together, these results show that chondroitin sulfate is involved in palatal wound healing.

Ductal carcinoma in situ with spindle cells: a potential diagnostic pitfall in the evaluation of breast lesions.

AIMS: To evaluate the morphological features of 11 cases of breast ductal carcinoma in situ (DCIS) with spindle cells and to propose an approach to distinguish it from benign mimics. The association with neuroendocrine differentiation was also investigated. METHODS: Cases of breast DCIS with a spindle cell component diagnosed in the Department of Pathology, Singapore General Hospital, between June 1996 and January 2003, were included in the study. The histological characteristics were documented, and immunohistochemistry for neuroendocrine markers, hormone receptors, cerbB2, smooth muscle actin (SMA) and high-molecular-weight (HMW) cytokeratins, was carried out. Electron microscopy was carried out on reprocessed paraffin-embedded material in three cases. RESULTS: Of 11 women diagnosed with DCIS with spindle cells, four presented with nipple discharge, six with a breast lump, while one was discovered to have a screen detected density. The tumour size ranged from 3 to 41 mm. The proportion of spindle cells varied from 10% to 80% of the in-situ tumour cell population. Nuclear grade was low in seven cases and intermediate in four. Necrosis was observed in two cases. Architectural pattern was papillary in six cases, and mixed in the rest. Microinvasion was present in two cases, with possible microinvasion in another two. Immunohistochemistry for neuroendocrine markers synaptophysin and chromogranin showed positive reactivity for at least one marker in all but three cases; one of these latter cases demonstrated ultrastructural neurosecretory granules. Oestrogen and progesterone receptors were expressed in 10 and nine cases, respectively, while cerbB2 was positive in only one case. HMW cytokeratin immunoprofile revealed a general lack of immunostaining within the abnormal cell population; likewise, no positivity for SMA of the cellular proliferation was detected. CONCLUSIONS: Almost all DCIS lesions with spindle cells disclose neuroendocrine differentiation. Although the distinction from benign florid usual hyperplasia may pose a diagnostic histological problem, the presence of diffuse neuroendocrine expression, in conjunction with the pattern of HMW keratin profile on immunohistochemistry, supports an in-situ neoplastic process. The absence of SMA immunostaining, in conjunction with negative reactivity for cytokeratins 5/6 and 14, makes the possibility of a myoepithelial proliferation unlikely.

Lactacystin-induced apoptosis of cultured mouse cortical neurons is associated with accumulation of PTEN in the detergent-resistant membrane fraction.

The tumor suppressor function of PTEN is attributed to its phospholipid phosphatase activity that dephosphorylates the plasma membrane phosphatidylinositol-(3,4,5)-triphosphate [PtdIns(3,4,5)P3]. Implicit in this notion is that PTEN needs to be targeted to the plasma membrane to dephosphorylate PtdIns(3,4,5)P3. However, the recruitment of PTEN to the plasma membrane is not fully understood. Here, we demonstrate PTEN accumulation in the detergent-insoluble fraction of neuronal cells in response to treatment by the proteasome inhibitor lactacystin. First, lactacystin induces apoptosis and the activation of caspase-3 in cultured cortical neurons. Second, PTEN undergoes proteolysis to form a truncated 50-kDa form that lacks parts of its C-terminal tail. Third, the truncated PTEN is stably associated with the detergent-insoluble fraction in which the plasma membrane marker protein flotillin-1 resides. Taken together, our results suggest that truncation and accumulation of PTEN to the detergent-insoluble membrane fraction are two events associated with the apoptotic signals of the proteasome inhibitor in cortical neurons.

The HercepTest and routine C-erbB2 immunohistochemistry in breast cancer: any difference?

INTRODUCTION: The proto-oncogene c-erbB2, located on chromosome 17q21, encodes a 185-kD transmembrane glycoprotein. It is known to be overexpressed, amplified, or both in 20% to 30% of breast cancers. C-erbB2 belongs to the human epidermal growth factor receptor (tyrosine kinase receptor) family that plays an important role in cell cycle regulation and differentiation. Although there are various methods to assess c-erbB2 status in breast cancer, protein overexpression determined by immunohistochemistry and gene amplification using fluorescence in situ hybridisation are most commonly utilised. This study compares the results of the DAKO HercepTest with the immunohistochemical assay (A0485, DAKO), which is routinely used in our pathology laboratory. MATERIALS AND METHODS: Paraffin-embedded breast cancer tissues from 41 patients operated in a tertiary hospital during the year 2000 were subjected to immunohistochemistry by the above methods. C-erbB2 positivity was defined by cytoplasmic membrane staining of 2+ or 3+ intensity. RESULTS: Overexpression of c-erbB2 protein was present in 36.6% and 41.5% of cases when detected by HercepTest and the DAKO A0485 antibody, respectively. There was almost perfect agreement between both methods (k = 0.898) when positive versus negative results were considered, and moderate agreement in terms of individual staining intensities (k = 0.554). CONCLUSION: Routine immunohistochemistry using the DAKO A0485 antibody is a reliable, cost-effective alternative to the HercepTest in determining prognosis and suitability of patients for Herceptin therapy.

Photoactivation of hypericin down-regulates glutathione S-transferase activity in nasopharyngeal cancer cells.

Photodynamic therapy (PDT) is a new modality of treatment for cancer. Hypericin is a photosensitizer, which is known to generate reactive oxygen species upon activation with light. We observed that photoactivated hypericin induces the generation of reactive oxygen intermediates in nasopharyngeal cancer (NPC) cells in vitro. There was also significant reduction of Glutathione S-transferase (GST) activity in HK1 and CNE-2 NPC cells and in tumor tissues from the NPC/HK1 murine tumor model by hypericin-mediated PDT. As antioxidants protect cells against phototoxicity, down-regulation of GST activity would potentiate the efficacy of hypericin-PDT treatment.

Identification of RANKL in osteolytic lesions of the facial skeleton.

RANKL (receptor activator of nuclear factor kappaB ligand) promotes osteoclast differentiation, stimulates osteoclast activity, and prolongs osteoclast survival and adherence to bone. Abnormalities of the RANKL/RANK/osteoprotegerin system have been implicated in a range of diseases, including osteoporosis. To date, no work has been done in osteolytic lesions of the facial skeleton. In this study, specimens of ameloblastomas, dentigerous cysts, odontogenic keratocysts, and radicular cysts were subjected to immunohistochemical analysis for RANKL and tartrate-resistant acid phosphatase (TRAP). Immunofluorescence staining for TRAP was visualized under confocal microscopy. All specimens demonstrated distinct positive immunoreactivity to RANKL and TRAP. The TRAP-positive cells also stained with in situ hybridization for human calcitonin receptor, a definitive marker for osteoclasts. Mononuclear pre-osteoclasts were observed to migrate from blood to the connective tissue stroma and multinucleate toward the bone surface. It can be concluded that RANKL plays a role in bone resorption in osteolytic lesions of the facial skeleton.

Keloids - the sebum hypothesis revisited.

The aetiology of the keloid scar has not been completely elucidated. Numerous hypotheses have been proposed in the past to explain the unusual characteristics of the keloid scar. While we do know that there is excessive and ongoing collagen-deposition, the exact triggering stimulus is a subject of conjecture. We present some of our photographic records of keloids and electron microscopic findings of keloid edges and reiterate the sebum hypothesis. We also attempt to explain the features of keloids in the light of the present knowledge of immunology and cell biology.

Correlation of nuclear morphometry with pathologic parameters in ductal carcinoma in situ of the breast.

Morphometric features of nuclear perimeter, nuclear area, feret ratio, and feret circle were studied in a series of 64 cases of ductal carcinoma in situ (DCIS) of the breast in Singapore women. The results were compared with pathologic parameters of tumor size, nuclear grade, necrosis, cell polarization, and architectural pattern. There was statistically significant correlation between nuclear perimeter and area with all the pathologic parameters, with the strongest association observed for nuclear grade (P <.0001). Higher grade nuclei as assessed histologically were associated with larger nuclear area (44.14 microm(2) in low-grade lesions, 47.77 microm(2) in intermediate-grade lesions, and 72.05 microm(2) in high-grade lesions) and perimeter (25.94 microm in low-grade nuclei, 27.12 microm in intermediate-grade nuclei, and 33.66 microm in high-grade nuclei). DCIS lesions with necrosis and absence of polarization also revealed increased nuclear area and perimeter (P <.05). Comedo architecture was associated with larger nuclear area and perimeter (65.97 microm(2), 31.7 microm) than the papillary subtype (42.17 microm(2), 25.29 microm), with the mixed morphologic pattern disclosing intermediate values (54.83 microm(2), 29.43 microm). There was direct correlation for tumor size with nuclear area and perimeter (P <.01). No similar relationship was found between pathologic parameters and feret ratio or circle, indicating that nuclear roundness or lack of it did not factor as a significant component in the pathologic assessment.

Metallothionein 1F mRNA expression correlates with histological grade in breast carcinoma.

Immunohistochemical expression of metallothioneins (MTs), a group of intracellular metal-binding proteins, is well documented in breast cancer. However, there is a paucity of information on the expression of the different MT isoforms in breast cancer tissues. The dichotomous association of MT overexpression with tumour types and progression led us to examine the role of the MT-1F mRNA isoform in breast cancer. We evaluated MT expression in 48 primary invasive ductal breast cancer tissues by immunohistochemistry, and the corresponding MT-1F mRNA expression via a semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) assay. The specificity of the RT-PCR products was confirmed by direct cycle sequencing and restriction enzyme digestion. Immunohistochemical analysis of MT revealed a significantly higher MT expression in histological grade 3 tumours as compared to grade 1 and 2 tumours (p = 0.021). Similarly, MT-1F mRNA expression was found to be significantly higher in grade 3 tumours (p < 0.001). The results suggest that the MT-1F isoform influences histological differentiation in invasive ductal breast cancer. The converse is also true in that the histological grade may determine the level of MT-1F expression in breast cancer.