BRCA1 and BRCA2 mutation testing in Cyprus; a population based study.

This paper presents the largest study in Cyprus evaluating the frequency and distribution of BRCA1/2 mutations in a high risk patient cohort. Deleterious mutations in the BRCA1/2 genes were identified in 68 of the 527 patients tested (13%). It is of interest that a quarter of those tested positive, did not have an extensive family history of breast/ovarian cancer but were diagnosed with early onset breast cancer, ovarian cancer under the age of 60 or triple negative breast cancer. The spectrum of mutations identified in our patient cohort is different compared to other Mediterranean countries. Furthermore, several of the mutations detected are novel and have not been identified in other ethnic populations. This highlights the importance of operating a national reference center for cancer genetic diagnosis which offers services tailored to the needs of the Cypriot population.

Marine and terrestrial foods as a source of brain-selective nutrients for early modern humans in the southwestern Cape, South Africa.

Many attempts have been made to define and reconstruct the most plausible ecological and dietary niche of the earliest members of the human species. While earlier models emphasise big-game hunting in terrestrial, largely savannah environments, more recent scenarios consider the role of marine and aquatic foods as a source of polyunsaturated fatty acids (PUFA) and other brain-selective nutrients. Along the coast of southern Africa, there appears to be an association between the emergence of anatomically modern humans and accumulation of some of the earliest shell middens during the Middle Stone Age (200-40 ka). Fragmentary fossil remains classified as those of anatomically modern humans, along with marine food residues and numerous material cultural indicators of increased social and behavioural complexity have been recovered from coastal sites. In this paper, new information on the nutrient content of marine and terrestrial foods available to early modern humans in the southwestern Cape is presented and compared with existing data on the nutritional value of some wild plant and animal foods in Africa. The results suggest that coastal foraging, particularly the collection of abundant and predictable marine molluscs, would have allowed early modern humans to exploit some of the richest and most accessible sources of protein, micronutrients and longer-chain omega-6 and omega-3 fatty acids. Reliable and accessible sources of omega-3 eicosapentaenoic and docosahexaenoic acid are considerably more restricted in terrestrial foods.

Early treatment with glucocorticoids or cyclophosphamide retains the slit diaphragm proteins nephrin and podocin in experimental lupus nephritis.

Renal podocytes and their slit diaphragms ensure the integrity of renal basement membrane and prevent urinary protein loss. We have previously reported that decreases of the podocyte slit diaphragm proteins nephrin and podocin represent early events in the podocytopathy of lupus nephritis (LN). We asked whether immunosuppressive agents such as glucocorticoids and cyclophosphamide may have direct effects on podocytes. We assessed in New Zealand Black/New Zealand White (NZB/W) F1 LN mice glomerular nephrin and podocin expression and localization by the use of Western blot and immunofluorescence; mRNA levels were measured by real-time polymerase chain reaction (PCR) and renal histology by light and electron microscopy. Early treatment with glucocorticoids and cyclophosphamide halted the histologic alterations associated with LN, preserving podocyte foot processes. Nephrin and podocin protein expression significantly increased in both glucocorticoid and cyclophosphamide groups as early as after three months of therapy. Real-time PCR revealed similar enhancement in nephrin and podocin mRNA levels after three to six months of treatment. This study documents that early treatment in experimental LN with glucocorticoids or cyclophosphamide preserves slit diaphragm proteins in podocytes and halts histological changes of the glomeruli, thus raising the possibility of a direct protective effect of these drugs on podocytes.

Podocyte main slit diaphragm proteins, nephrin and podocin, are affected at early stages of lupus nephritis and correlate with disease histology.

Renal podocytes and their slit diaphragms ensure the integrity of the renal basement membrane that forms the barrier to urinary protein loss. A putative disruption of the slit diaphragm and its main protein components, nephrin and podocin, may be implicated in the pathogenesis of lupus nephritis (LN). We studied the glomerular protein expression of nephrin and podocin in NZB/W LN mice by Western blot and immunofluorescence; mRNA levels were measured by real-time PCR. Human kidney biopsies of class II (n = 5), IV (n = 4), V (n = 7) LN were evaluated for nephrin expression by immunohistochemistry. Glomerular protein expression of nephrin and podocin were significantly reduced in NZB/W LN, starting from the earlier stages (mild mesangial LN) and becoming pronounced at advanced histological forms (focal and diffuse proliferative LN). Nephrin and podocin mRNA levels were substantially decreased in diffuse proliferative disease. Decreased expression of both proteins correlated with electron microscopy findings of distorted slit diaphragms. In patients with LN, nephrin was decreased particularly in diffuse proliferative LN. The main slit diaphragm proteins, nephrin and podocin, are affected from the earlier stages of LN and their expression correlates with disease histology. Our findings suggest a novel role of podocytes and their structures in immune-mediated nephritis.

Human muscle-derived stem cells. Effectiveness in animal models of faecal incontinence. Research scheduling.

Researchers believe that human muscle-derived cells are able to restore leak-point pressure to normal levels by differentiating into new muscle fibres that prevent anal sphincter muscle atrophy. Laboratory data are needed to identify exactly how these cells work to regenerate muscle. The objective of this study is to test whether stem cells can be employed to treat internal anal sphincter (IAS) injuries in humans; to this end, this work will use a two-step process to study: first, the effectiveness of the treatment in a sample of animals with artificial injuries to the IAS and then to verify the results in a population of selected humans affected by pathology.

Thyroid tissue remnants after "total thyroidectomy".

Total Thyroidectomy (TT) is a gold standard for benign bilateral pathologies and malignant pathologies of the thyroid. TT has numerous advantages over less radical approaches, such as the resolution of the thyroid pathology, avoidance of recurrences, and improved response to life-long substitutive organotherapy. TT has a negligible rate of recurrence. Near Total Thyroidectomy (NTT) is associated with a low rate of recurrence. Subtotal Thyroidectomy (ST), in which a portion of the thyroid gland is deliberately left in the thyroid lodge, has a considerably higher rate of recurrence. The incidence of complications with TT is similar to that with other techniques of thyroid exeresis. However, despite the radical intent of surgeons, a real TT is not always carried out. The complete removal of all the thyroid tissue employing TT is not the norm and micro/macroscopic remnants almost always remain. The literature on these tissue remnants is often based on techniques that are not very accurate in terms of determining the diameters of the tissue remaining. In our study, conducted by colour echo-doppler of the thyroid lodge in 102 patients who had undergone TT for benign thyroid pathologies, we demonstrated significant thyroid tissue remnants after TT in 34 cases of 102 (33,3%). Therefore, out of a total of 102 so-called "total thyroidectomies", only 68 (66,7%) were really total, whereas 12 patients (11,76%) had near total thyroidectomy, leaving tissue remnants < 1 cm, and 22 patients (21,57%) had subtotal thyroidectomy, with tissue remnants > or = 1 cm.

Mitochondrial encephalomyopathies: a review of routine morphological diagnostic methods with emphasis on the role of electron microscopy.

Mitochondrial encephalomyopathies (MEs) are a group of clinically and genetically heterogeneous diseases. They can be caused by defects in both mitochondrial or nuclear coded genes. Their phenotypic expression is governed by unique biological phenomena such as the dual genetic control, mitotic segregation, heteroplasmy and threshold effects. Currently, the correct diagnosis of ME relies on a multidisciplinary approach which includes clinical information as well as laboratory data from muscle morphology, biochemistry and molecular genetics. Among the morphological methods, histology, histochemistry and electron microscopy were historically instrumental in the diagnosis of MEs. However, with the development of molecular genetics, the diagnostic value of morphology and of electron microscopy in particular have been questioned. The aim of the present review is to present a comparative assessment of the diagnostic contribution of histology, histochemistry and electron microscopy in a group of 48 patients with a diagnosis of ME.

Medullary cystic kidney disease with hyperuricemia and gout in a large Cypriot family: no allelism with nephronophthisis type 1.

We describe a large Cypriot family with an interstitial type of nephropathy, inherited as an autosomal dominant trait that led to end stage renal failure between 51 to 78 years of age (mean 62.2 years). Twenty-three people are known to be affected, but several younger relatives with normal renal function may remain undiagnosed because of the absence of precise clinical and laboratory diagnostic criteria. This nephropathy is associated with medullary renal cysts, hypertension, hyperuricemia, and gout. Several relatives have typical medullary cystic disease (MCD), while in the others the findings are compatible with this diagnosis. Due to the similarity of clinical and pathologic findings, earlier reports had suggested that MCD may be allelic to autosomal recessive familial juvenile nephronophthisis, which was mapped recently to chromosome band 2q13. Linkage analysis of the present family with a closely linked marker excluded linkage to the above locus. Linkage was also excluded to the PKD1 locus of adult polycystic kidney disease type 1, and up to 5 cM on either side, on chromosome 16. We suggest that because of the element of hyperuricemia and gout found in this family, although with reduced penetrance, it may represent a variant of autosomal dominant MCD of the adult type. This variability may be the result of allelic or locus heterogeneity. Molecular genetic approaches including linkage analysis on appropriate families will certainly assist in classifying such related genetically heterogeneous disorders.

Distal hereditary motor neuronopathy of the Jerash type.

A novel form of autosomal recessive distal hereditary motor neuronopathy (distal HMN) is reported. The presence of pyramidal signs within the early stages of the disease with persistence of knee hyperreflexia form distinctive clinical features. We have mapped the HMN-J gene to chromosome 9p21.1-p12, within an estimated interval of 1.2-Mb.

Unusual skin tumors in Langerhans' cell histiocytosis.

A case of Langerhans' cell histiocytosis with unusual skin manifestations in the form of multiple large skin tumors is described. The skin lesions responded partially to chemotherapy with etoposide and prednisone, and residual lesions were excised surgically. The patient developed central diabetes insipidus during treatment.

AM-FM texture segmentation in electron microscopic muscle imaging.

This paper describes the application of an amplitude modulation-frequency modulation (AM-FM) image representation in segmenting electron micrographs of skeletal muscle for the recognition of: 1) normal sarcomere ultrastructural pattern and 2) abnormal regions that occur in sarcomeres in various myopathies. A total of 26 electron micrographs from different myopathies were used for this study. It is shown that the AM-FM image representation can identify normal repetitive structures and sarcomeres, with a good degree of accuracy. This system can also detect abnormalities in sarcomeres which alter the normal regular pattern, as seen in muscle pathology, with a recognition accuracy of 75%-84% as compared to a human expert.

BRCA1 germline mutations in Cypriot breast cancer patients from 26 families with family history.

Germline mutations in the BRCA1 gene are causative for a variable number of hereditary breast/ovarian cancers. The data presented in this study are based on genetic analysis of the BRCA1 gene in 49 DNA samples from breast cancer patients with a positive family history. A combination of manual direct DNA sequencing and SSCP analysis was used to screen the entire coding region of BRCA1. Overall 13 variants were detected which included 5 missense mutations, 3 polymorphisms and 5 intronic changes. Further genetic analysis of the 13 variants was carried out using 50 control DNA samples. Our results showed that 12 out of the 13 variants detected in the DNA of the patients group, were also present in the control group. It appears that the Greek Cypriot families studied so far have an unexpectebly low frequency of deleterious mutations in the BRCA1 gene. This is the first report on BRCA1 mutation analysis in Cyprus.

A comparative morphological study in 33 cases of respiratory chain encephalomyopathies.

Mitochondrial encephalomyopathies (ME) are clinically and genetically heterogeneous syndromes ranging from pure myopathies to complex multisystem disorders. This phenotypic and genotypic variability, coupled with the lack of a laboratory gold standard marker for the diseases, makes diagnosis a challenging process. Mitochondrial DNA analysis and biochemical assay of muscle homogenates are quite specific diagnostically but have low sensitivity in unselected cases suspected of ME. We decided to evaluate four routine morphological methods in 33 cases of definite or probable ME in an effort to assess the reliability of each of these techniques in diagnosing ME.

Morphological changes in the rabbit submandibular gland after parasympathetic or sympathetic denervation.

Selective denervation was used to obtain further information about the relative roles of the nerves supplying the gland. Parasympathetic denervation soon caused atrophy in both acinar and granular tubule cells, and a substantial reduction in gland wet weight. The secretory cells had several unusual features, and striated duct cells tended to accumulate glycogen. Three to four weeks after pre-ganglionic parasympathectomy, the parenchymal cells were less abnormal but still smaller than in the control, unoperated gland. Thus, parasympathetic impulses are required to maintain normal parenchymal cells. Chronic sympathectomy caused little or no detectable changes in the secretory cells, and there was no significant change in wet weight. Nevertheless, pre-ganglionic sympathetic axotomy reduced the reflex secretion of acinar mucosubstances in response to feeding. Thus, sympathetic impulses normally contribute to the reflex secretion of acinar mucosubstance from these glands.

Structural and functional studies of the effects of sympathetic nerve stimulation on rabbit submandibular salivary glands.

Continuous sympathetic stimulation at 8-10 Hz caused intense vasoconstriction in the gland, so stimulation was generally given in an interrupted pattern to minimize this detrimental effect on secretion. Only a small increase in fluid secretion occurred; it became thick and tended to block the cannula; therefore in later experiments the main duct was not cannulated. After sympathetic stimulation there was substantial degranulation of acinar cells. However, as this was accompanied by little movement of water, the secreted mucosubstance distended the ductal lumina. The granular tubule cells were unchanged by sympathetic stimulation. Use of selective blocking agents revealed that the sympathetically-evoked secretion of acinar mucin was mediated mainly via beta-adrenoreceptor activation. As stimulation of the sympathetic nerves alone caused little additional formation of fluid, the effects of superimposing continuous low frequency sympathetic stimulation onto a background of low parasympathetic secretion were compared with similar parasympathetic stimulation alone of the contralateral gland. These double nerve stimulations did not augment the volume of fluid secreted, or cause morphological changes additional to those from parasympathetic stimulation alone. Nevertheless, it is likely that, under natural reflex conditions, sympathetic impulses can increase the amount of acinar mucosubstance secreted.

Structural and functional studies of the effects of parasympathetic nerve stimulation on rabbit submandibular salivary glands.

The effects were assessed by combining physiological experiments with morphological examination, using light and electron microscopy. Continuous, parasympathetic nerve stimulation, at frequencies varying from 1 to 10 Hz, caused a copious flow of saliva. Both acinar and granular tubule cells showed extensive degranulation, but the effects on tubule cells were the more dramatic. After stimulation, there was a marked loss of acidic mucosubstances from the acinar cells and an almost complete loss of neutral mucosubstances from the granular tubule cells; this was particularly evident with stimulation at higher frequencies (6-10 Hz).

Amyloid myopathy: evidence for mechanical injury to the sarcolemma.

Myopathy is a rare clinical manifestation in primary systemic amyloidosis. The clinical phenotype and muscle histology are well described but the pathophysiological mechanisms remain poorly understood. We report a 40-year-old man who presented with hypertrophic cardiomyopathy and a limb girdle syndrome associated with deposition of amyloid and free lambda light chains in skeletal muscle. Electron microscopy showed amyloid fibrils, physically disrupting the plasma membrane and basal lamina, while laminin immunocytochemistry revealed a reduction of laminin beta1 and upregulation of laminin alpha1. We believe that one of the possible pathophysiological mechanisms in amyloid myopathy is mechanical disruption of the sarcolemma by the abutting amyloid fibrils.

Computer-aided detection of breast cancer nuclei.

A computer-aided detection system for tissue cell nuclei in histological sections is introduced and validated as part of the Biopsy Analysis Support System (BASS). Cell nuclei are selectively stained with monoclonal antibodies, such as the anti-estrogen receptor antibodies, which are widely applied as part of assessing patient prognosis in breast cancer. The detection system uses a receptive field filter to enhance negatively and positively stained cell nuclei and a squashing function to label each pixel value as belonging to the background or a nucleus. In this study, the detection system assessed all biopsies in an automated fashion. Detection and classification of individual nuclei as well as biopsy grading performance was shown to be promising as compared to that of two experts. Sensitivity and positive predictive value were measured to be 83% and 67.4%, respectively. One major advantage of BASS stems from the fact that the system simulates the assessment procedures routinely employed by human experts; thus it can be used as an additional independent expert. Moreover, the system allows the efficient accumulation of data from large numbers of nuclei in a short time span. Therefore, the potential for accurate quantitative assessments is increased and a platform for more standardized evaluations is provided.