Understanding Disease Pathogenesis

Cystic fibrosis illustrates how our understanding of this disorder has changed through molecular genetics. What was previously a ''disease'' is now a syndrome because DNA-based knowledge has shown that a number of sporadic conditions are part of the cystic fibrosis spectrum. These include meconium ileus in the fetus, male infertility related to congenital bilateral absence of vas deferens (CBAVD), some forms of disseminated bronchiectasis, and chronic idiopathic pancreatitis. Evidence that the...

Clinical Applications

Denaturing gradient gel electrophoresis can be used for genetic diagnosis of hereditary human diseases caused by small mutations even when doing diagnosis for X-linked human diseases. It is also possible to analyze SNPs (genetic alterations where at least one allele frequency is higher than 1 ) to associate with hereditary complex disease traits. In many cases, mutation detection can be used in choosing adequate healthcare, as some mutation positions are associated with the worst prognostic....

Snp Genotyping

High-throughput SNP genotyping methods have been developed in order to exploit the many healthcare benefits which will arise from a detailed knowledge of genetic variation. 1-3 SNP genotyping technologies have two components a method for determining the type of base present at a given SNP locus (allele discrimination), and a method for reporting the presence of the allele(s) (signal detection). 4-6 There are three general allele discrimination methods hybridization annealing (with or without a...

Doublecontrolled Conditionally Replicating Adenoviruses

After the systemic administration of adenoviral vectors, most of the virus ends up in the liver. When adenoviral replication is not strictly restricted to certain cell types or tissues, severe liver damage might occur due to adenoviral replication and consequential lysis of the liver cells. Because of this importance to restrict adenoviral replication, solutions have to be found for the observed aspecific replication of CRAds. One solution is a double-controlled conditionally replicating...

Clinical Description

The syndrome was initially described as the association of five major features paucity of interlobular bile ducts, peripheral pulmonary artery stenosis, butterfly-like vertebral arch defect, posterior embryotoxon, and a peculiar facies. 1 However, minor features such as renal abnormalities, skeletal defect, high-pitched voice, and mental and growth retardation were also described. In addition, some patients occasionally present with clinical features including other arterial stenosis, small...

C1494T Mutation In The 12S rRna Gene

The homoplasmic C1494T mutation was recently identified in a large Chinese family with AID and maternal inheritance.1-14-1 Twenty of 39 matrilineal relatives, who are supposed to carry the mutation, exhibited hearing loss. Clinical data have shown that the treatment with amino-glycosides can induce or worsen deafness in matrilineal relatives. The mutation occurs at a position where the A at position 1555 can make a pair with the mutant base (Fig. 1D). Additional data indicated that the cell...

Fetal Cells Diagnosis Or Screening

The scarcity of fetal cells in the maternal circulation and the failure to reliably retrieve these cells from maternal blood samples are amply underscored by the sobering results of the NIFTY Study. As even more disappointing results were obtained in the study initiated by Applied Imaging, which used state-of-the-art technology for the automated detection of putative fetal erythroblasts, 5 questions have been raised as to whether the pursuit of fetal cells can have any future diagnostic value....

Pcrldr And GapLCR

A major problem of the LCR reaction is joining of blunt end duplexes to each other and to themselves, all of which can serve as templates for further amplification, although no real target DNA is present in the original sample 7 and this poses significant limitations on the sensitivity of LCR. 6 To overcome this problem, several modifications have been developed, including a combined PCR LDR method and Gap-LCR (G-LCR). In PCR LDR, the amplification step for LDR is PCR utilizing outside primers...

Comparison Of Viral Load Assays

A number of studies have compared the performance of these assays in clinical situations. Schuurman et al. 4 showed that interlaboratory reproducibility of the Roche, Organon, and Versant assays was very good. Others have shown that each of the assays has relatively low intra- and interassay variability.1-5-1 The biological variation of HIV-1 RNA in plasma in clinically stable individuals is approximately threefold (0.5 log) and this variation was not associated with diurnal fluctuations.1-6-1...

Pharmacogenetics

Much of the individual variation in drug response is due to genetic polymorphisms of drug-metabolizing enzymes, and genomic testing for drug-metabolizing enzymes has significant potential for improving the efficiency of drug treatment and reducing adverse drug reactions and thus promises to improve future health care in a number of ways. It is very likely that these tests will make up a significant proportion of total nucleic acid-based testing in the coming years. 23 Currently, however, only a...

Therapeutical Approaches for HCM Are Symptom Oriented

P-Blockers act as negative inotropic drugs and are used as first-line drugs in patients with or without LV obstruction, and are the preferred treatment in symptomatic HCM patients with an LV outflow tract gradient only by exertion. 5 Verapamil, a calcium antagonist with negative inotropic properties, is used in both obstructive and nonobstructive forms in patients without severely disabling symptoms. Disopyramide, a class IA antiarrhyth-mic drug with negative inotropic effects, is reserved for...

Materials and Methods

Dilute 50-500 ng of target DNA to a final volume of 5 mL with 10 mM Tris-HCl (pH 8.5), -0.1 mM EDTA and denature the DNA for 5 min at 98 C in a thermocycler with heated lid. Mix 1.5 mL probe mix (1 fmol of each probe) and 1.5 mL MLPA buffer (1.5 M KCl, 300 mM Tris-HCl pH 8.5, and 1 mM EDTA) and add to the denatured DNA. Heat the mix for 1 min at 95 C before incubating overnight (16 h) at 60 C. After hybridization dilute the sample to 40 mL with ligase mix (2.6 mM MgCl2, 5 mM Tris-HCl pH 8.5,...

Pcr Inhibition Or Contamination

The use of an internal control is important because impurities or improper nucleic acid concentration may inhibit the PCR reaction, resulting in false negatives. The internal control will give a greater degree of confidence in the validity of a negative amplification result. Every amplification test where inhibitors such as nontarget DNA, heme, bilirubin, bile salts, ethylenediaminetetra-acetic acid (EDTA), sodium dodecyl sulfate (SDS), proteases, and bacterial extracts, etc. are problematic...

Psychiatric Disorders

Upon treatment with antipsychotic drugs, adverse reactions are more likely to be seen in PMs for CYP2D6. In addition, the costs for treatment of patients are higher and estimates have been made that costs 4000-6000 more per year to treat patients of the variant UM and PM phenotypes. 9 Parkinsonism-like side effects are Relative importance of polymorphisms in human P450s involved in drug metabolism

Genomics And Molecular Typing

The Chlamydia genome is composed of a chromosome of approximately 1.04 million base pairs and a plasmid of 7493 base pairs. 19 The trachoma and LGV biovars have high gene sequence homology and usually only single base-pair mutations are found between the two. Several genes show heterogeneity that is important. The major outer membrane protein (MOMP) gene exhibits extensive sequence variation within the serovars. The differences are mainly clustered in four regions (VD1 through 4), which form...

The Neurological Continuum

Although type I is, by definition, nonneuronopathic, and the presence of the N370S allele obviates neurological involvement, nonetheless, recent data are suggestive of a phenotypical continuum that includes some neurological signs even in patients with type I. Among the earliest anecdotal reports were those of parkinsonism that is marked by early onset, aggressive progression, and refractoriness to conventional anti-Parkinson therapy in patients with very mild type I Gaucher disease. 6...

Description Of The Genus Taxonomy

C. pneumoniae belongs to the genus Chlamydia, which also contains the species Chlamydia trachomatis, Chlamydia psittaci, and Chlamydia pecorum. Due to the detection of new Chlamydia-like organisms and genetic analysis, in 1999 a new taxonomy was proposed which divides the common chlamydiae into the two families, Chlamydophila (including C. pneumoniae) and Chlamydia, and adds two more families containing the newly detected chlamydia-like organisms (Table 1). 1 This proposal, however, is still...

Extracellular Circulatory Fetal

Possibly the observation that has changed future prospects for the noninvasive diagnosis of fetal genetic traits the most was that describing the presence of extracellular fetal DNA in maternal plasma and serum. 8 This analysis was prompted by reports indicating that cell-free tumor-derived DNA was readily detectable in the plasma of cancer patients. 9 By arguing that the placenta shares many similarities with tumors (high rate of cell turnover, tissue invasion), it was hypothesized that...

Virulence Factors

In human experimental models, bacteria are delivered into the skin via puncture wounds using an allergy-testing device. 3 An estimated delivered dose (EDD) of 1 colony-forming unit already results in 50 papule formation. 4 Most bacteria localize in the dermis after 48 hr of infection. Several potential H. ducreyi virulence factors have been reported in the literature and their possible effects were studied using isogenic mutants by grafting both wild-type strains (i.e., 35000 HP) and mutant...

Genes And Parkinsonism

Parkinsonism is a syndrome of rest tremor, rigidity, bradykinesia, loss of postural reflexes, and the freezing phenomenon.1-1-1 Most often, it is a manifestation of Parkinson's disease (PD), which is characterized neuro-pathologically by brainstem Lewy bodies. Recent epide-miological and molecular data suggest that this is not one disorder, but is comprised of at least several disorders with genetic, environmental, or mixed etiologies. Genes play a complex role in PD. Most twin studies, which...

Laboratory Diagnosis

In clinical practice, e.g., for the planning of operations, diagnosis of congenital F XI deficiency is usually established by APTT-based assays, whereas molecular techniques are mostly restricted to scientific studies. However, there are conditions, such as consumption reactions or the presence of a high titer of lupus anticoagulant, where APTT-based tests may not be suitable and molecular methods are needed to identify a congenital deficiency state. For mutation analysis, genomic DNA is...

Clinically Relevant Pilot Applications

The practical RCA potential to identify nucleic acid targets, antibodies, and antigens in clinical samples has recently been demonstrated in several feasibility studies. Specifically, the RCA-based protocol for an automated scoring of single nucleotide polymorphisms (SNPs) in a set of human genomic DNA samples with the nanogram sensitivity was developed. 15,20 Highly sensitive multiplex detection of hotspot somatic mutations present at very low abundance was also reported. 21 The RCA capability...

Simultaneous Detection Of Several Mollicute Species

As mollicutes usually cause diseases of the respiratory or the urogenital tract, where the spectrum of pathogens to be considered is large, considerable interest has been focused on the simultaneous detection of several molli-cutes, or mollicutes and other pathogens, by the same assay. Although the regions V2 and V3 of the 16S rDNA display sufficient interspecies differences for species-specific detection, conserved sequences of the 16S rDNA gene enable detection of most Mycoplasma spp.,...

Other Mutations In The 12S rRna Gene

Deletion of a T at position 961 and insertion of varying numbers of G were observed in a patient during a search in the mtDNA in 35 sporadic Chinese cases with AID. 30 The same mutation was not detected in 799 control samples. 30 This mutation was later found in an Italian family, in which five maternal relatives became deaf after aminoglycoside exposure. 31 However, in a recent study, 961delTinsC(n) was detected in 7 of 1173 anonymized blood samples in the United States. 24 This result...

Treatment and Control

Recent studies have shown that a single dose of azithromycin is as effective as a long course of topical tetracycline (42 doses over a 6-week period) in treating trachoma, and the effect of treatment lasts for 1 year. 3,4 However, only small differences in effectiveness have been found between mass treatment programs and programs treating active cases only. 5 WHO has set a goal of eliminating trachoma by 2020, using the SAFE program. 6,7 ''SAFE'' stands for surgery to correct lid deformities,...

Enteritis Necroticans Disease Summary

Enteritis necroticans (also known as necrotizing enteritis, Darmbrand, or Pigbel) caused by C. perfringens type C isolates is a rare but potentially lethal enteric disease of humans. This illness was first recognized after World War II in Darmbrand, Germany and later in Papua New Guinea. 20 The p -toxin produced by type C isolates is considered as the primary virulence factor based on the relative efficacy of a p-toxoid vaccine. 20 Although the incidence of enteritis necroticans is low, risk...

Oncolytic Adenoviruses

In the battle against cancer it is very important to kill every tumor cell present otherwise, the remaining malignant cells will continue to grow and form new tumors. Although adenoviral vectors are one of the most efficient vectors used, one injected dose is thought to infect maximally 10 of the tumor cells. Thus it is necessary to increase the efficiency of adenoviral infection. An approach to increase this efficiency is to allow the virus to replicate, thereby increasing the amount of...

Gsd Type Vi Liver Phosphorylase Hers Disease

The enzyme deficient in type VI GSD is liver-specific phosphorylase. It is relatively rare and a benign form of GSD with no heart and skeletal muscle involvement. Most patients with GSD VI disease present with hepatomegaly and growth retardation. Hypoglycemia, hyperlipidemia, and hyperketosis, if present, are usually mild. The hepatomegaly improves and disappears around puberty. Diagnosis rests on enzyme analysis of the liver biopsy. GSD VI is autosomal recessive. Liver phosphorylase gene...

Genotypephenotype Correlation

No correlations are found between a mutation and a clinical phenotype because a family unique presentation such as migraine or stroke is also found with several other mutations. 2,3 Additionally, we did not notice neither different clinical presentations nor different skin vessel wall lesions in CADASIL patients with mutations located either in the first or very last exons. Interestingly, a homozygous patient followed in Finland was indistinguishable when compared with an age-matched...

Gsd Type Xi Hepatic Glycogenoses With Renal Fanconibickel Syndrome

This rare GSD is caused by defects in the facilitative glucose transporter 2 (GLUT-2), which transports glucose in and out of hepatocytes, pancreatic cells, and the basolateral membranes of intestinal and renal epithelial cells. 30 The disease is characterized by proximal renal tubular dysfunction, impaired glucose and galactose utilization, and accumulation of glycogen in liver and kidney. The affected child presents in the first year of life with failure to thrive, rickets, hepato- and...

Thalassemia Major

The clinical picture of p-thalassemia major is characterized by presentation with pallor and spleen and liver enlargement at 6-24 months of age. Typical skeletal changes resulting from expansion of the bone marrow are evident only in patients in whom transfusions were initiated late or are undertransfused. At the age of 10-11 years, regularly transfused thalassemia major patients are at risk of developing severe complications related to iron overload. These complications include growth...

Differential Diagnosis

Most clinicians still rely on a set of criteria originally formulated by Kimonis et al. 1 Table 1 shows a current list of qualifying criteria. It is worth noting that a number of other related but apparently distinct disorders are recognized. Rombo syndrome 2 is a familial disorder with vermiculate atrophoderma, milia, hypotrichosis, trichoep-itheliomas, basal cell carcinomas, and peripheral vasodi-lation with cyanosis, whereas Bazex-Dupre-Christol syndrome 3 combines major characteristics of...

Introduction

Prader-Willi syndrome (PWS), a condition initially described by Prader, Labhard, and Willi in 1956, 1 is a neurodevelopmental disorder with a clinical presentation that includes neonatal hypotonia, obesity, mental retardation, and behavioral abnormalities. After introduction of high-resolution chromosome analysis, PWS was the first abnormal phenotype found to be associated with a submicroscopic chromosomal deletion. 2 The PWS-asso-ciated microdeletion spans about 4 Mbp in the chromosomal region...

Localized Skin and Soft Tissue Infection

Most species of NTM have been described causing cutaneous disease 9,10 (Table 2) or nosocomial skin and soft tissue infection, postinjection abscess, surgical wound infections, and infections because of long-term intravenous or peritoneal catheters. M. chelonae causes disseminated nodular skin disease in immunocompromised patients. M. marinum causing ''swimming pool granuloma'' or ''fish tank granuloma'' gets implanted while cleaning fish tanks or handling fish. Solitary papules get Table 1...

Clinical Features In Hereditary Breast And Ovarian Cancer Syndrome

BRCA mutations occur in approximately 20 of families with inherited susceptibility to breast cancer. Personal and family characteristics associated with an increased likelihood that a BRCA mutation will be identified are listed in Table 3. Breast cancers in BRCA1 carriers often have medullary features, are more likely to be poorly differentiated with high mitotic rates and S-phase fraction, lack an in situ component, have low estrogen and progesterone receptors score, and node-positive. BRCA2...

Screening In Hereditary Breast And Ovarian Cancer Syndrome

Although overall population screening reduces breast cancer mortality by 25-40 in women between the ages of 50 and 70 years, no data are available on the outcomes of interventions to reduce risk in HBOC syndrome. No data are available to demonstrate that surveillance for ovarian cancer in high-risk women reduces mortality 3 because transvaginal ultrasound and CA-125 lack sensitivity and specificity. However, recommended screening in BRCA gene mutation carriers includes monthly breast Table 4...

Substrate Reduction Therapy

Oral substrate reduction therapy (SRT) involves attenuation of the rate of synthesis to achieve a balance with reduced endogenous glucocerebrosidase. A pivotal trial of SRT was carried out in 28 adult patients (from four centers Cambridge, UK Amsterdam Prague and Jerusalem) naive to enzyme therapy, with mild to moderate type I Gaucher disease, who were unable or unwilling to receive enzyme treatment. There was significant reduction in spleen and liver, although amelioration of anemia and...

Bladder Cancer Studies Using In Vitro Models

Expression profiling using bladder cancer cell lines has been used to gain insight into the molecular events associated with clinical disease states, assigning potential functional roles to novel genes in both tumorigenic and tumor progression processes. Certain studies have focused on gene and pathway discovery associated to genistein 10 or 5-aza-2'-deoxycytidine 11 exposure. Other reports describe the functional classification of genes comparing the expression patterns of p53-mediated...

Molecular Basis And Genetic Counseling

PWS is the result of deficiency of one or more genes that map on the proximal long arm of chromosome 15 (Table 2). These genes are imprinted. Only their paternally derived alleles are functional.1-3-1 The genomic imprinting on chromosome 15 is reflected in the patterns of cytosine methylation in the CpG islands of this region. These cytosines are methylated only in the maternally derived sequences in the imprinted area. These detectable differences can be used for the diagnostic workup of...

Microarrays As Target Identification Tools In Cancer

Microarrays constitute a group of technologies characterized by the common availability of measuring hundreds or thousands of items, including DNA sequences, RNA transcripts, or proteins, within a single experiment using miniaturized devices. Hybridization-based methods and the microarray format constitute together an extremely versatile platform provided for both static and dynamic views of DNA structure, as well as RNA and protein expression patterns in cultured cancer cells and tumor

Prevalence And Clinical Diversity

OI is an autosomal genetic disorder characterized by brittleness of bones and susceptibility to bone fractures from very mild trauma. Fractures may be rare or frequent (more than 200 prior to puberty), and bone fragility may be accompanied by reduced life span and various connective tissue abnormalities short stature, blue sclerae, joint laxity, dentinogenesis imperfecta, easy bruising, and hearing loss. OI represents extreme variation in severity from lethal perinatal to barely detectable....

Udpglucuronosyltranferase 1a1

UDP-glucoronosyltransferases are enzymes responsible for detoxification and elimination of various metabolites and drugs. These enzymes also catalyze the inactivation of irinotecan (CPT-11), an analog of the alkaloid campto-thecin. CPT-11 is clinically used for the treatment of refractory and advanced colorectal cancers. Dose-limiting toxicities are diarrhea and pancytopenia. CPT-11 is metabolized to its active product, SN-38, by the enzyme carboxylesterase 2. As depicted in Fig. 3, UGT1A1 is...

The Need For Biomarkers In Bladder Cancer

Bladder tumors are pathologically stratified based on stage, grade, tumor size, presence of concomitant carcinoma in situ, and multicentricity.1-2-1 The chances of tumor progression are augmented with the increase of these pathological variables. Pathologically, most bladder tumors are transitional cell carcinomas. There is, however, increasing recognition of the prognostic importance associated with the metaplastic variants displaying squamous and glandular differentiation as part of their...

Chemoprevention In Hereditary Breast And Ovarian Cancer Syndrome

Women who desire risk reduction intervention for breast cancer have the option of chemoprevention with tamoxifen therapy (20 mg day for 5 years) or prophylactic surgery. The National Surgical Adjuvant Breast and Bowel Project prevention trial in healthy women 35 years and older with a 1.7 or greater cumulative 5-year risk for developing breast cancer demonstrated short-term reduction in risk of developing estrogen-receptor-positive breast cancer by 49 . The utility of tamoxifen for breast...

Pharmacogenetics Of Drugmetabolizing Enzymes

Polymorphism of genes encoding drug-metabolizing enzymes is the major known genetic cause for the interindividual variability in drug response. 1-6 Based on the occurrence of mutations in these genes, gene deletions, and gene duplications, the populations can be divided into poor (PM), intermediate (IM), efficient (EM), or ultrarapid (UM) drug metabolizers. The PMs lack the enzyme in question, the IMs are heterozygous for a defect gene, the EMs have two functional alleles, whereas the UMs have...

Bladder Cancer Studies Using Clinical Specimens

Microarray analyses have been used to correlate changes in the expression of specific genes and groups of genes within distinct bladder subclasses. Such signature genes would ideally provide a molecular basis for classification, yielding insight into the molecular events underlying different clinical bladder cancer phenotypes. The first report monitored the expression patterns of superficial and invasive tumor cell suspensions prepared from individuals and pools of normal and bladder tumors of...

Molecular Genetic Testing

Given the ever-growing list of CFTR defects, molecular genetic testing has evolved from relatively simple assays directed only at the predominant AF508 mutation 10 to increasingly complex multiplex mutation detection systems. Not all of these are on micro- or miniarray platforms, although the first practical assays for detecting more than a few mutations at a time utilized allele-specific DNA probes bound to solid supports in the form of nylon strips or membranes. 11,12 Originally all such...

Molecular Genetic Testing Genetic Counseling And Clinical Management

Molecular genetic testing in familial melanoma is currently considered premature by the International Melanoma Genetics Consortium1-23-1 for the following reasons. First, the mutation detection rate in melanoma families is very low. Second, risk estimates for mutation carriers are not well established (and have broad confidence intervals). In particular, the risk for other cancers, such as pancreatic cancer, is not clearly defined. Third, surveillance programs for mutation carriers have not yet...

Gender Bias

There appears to be a gender-specific influence on the degree and rate of disease progression in FSHD. The Fig. 3 The length of the smallest EcoRI fragment associated with the disease in unrelated FSHD individuals. Ten micrograms of DNA was digested with EcoRI, fractionated on a 0.5 agarose gel for 48 hr at 0.5 V cm, and Southern-blotted onto HybondN (Amersham). The DNA on the membrane was hybridized with radiolabeled DNA probe p13E-11. Lane 1 contains high-molecular-weight markers. Lanes 2-12...

Ethical And Societal Considerations

Although to date only symptomatic management can be recommended for type II disease, ethical issues relating to ERT that are neither life-prolonging nor appreciably improve the quality of life of these babies need to be addressed. Indeed, ethical arguments apply to the emotional outlay in neuronopathic forms, but also to the financial outlay for orphan drugs for nonsevere, non-neuronopathic patients. No evidence-based consensus has been documented to support the extraordinary expense of...

The Challenge Of Integrating Genomic Innovation

It is commonly believed that innovations succeed on their own merit. Those without real merit fail in the market place those with superior merit succeed simply on the basis of their superiority over existing alternatives. However, experience shows otherwise. Technological capability and superior engineering do not, in fact, guarantee acceptance (Table 1). Great innovations themselves are often insufficient to ensure usability, effectiveness, or user satisfaction. In other words, good technology...

Clinical Description And Prevalence

The incidence of melanoma is rising faster than all other cancers except lung cancer in women, currently varying between 5 (Western Europe) and 20 (Northern Europe) to over 50 (Queensland, Australia) cases per 100,000 per annum. 1,2 Familial clustering of melanoma was first described by Norris in 1820, 3 but it was not until the second half of the 20th century before others documented the familial occurrence of melanoma. Across several population-based studies, 1-13 of melanoma cases reported...

Famccggcccagccaccatggtamra

Table 3 Sequences of the probes used for real-time PCR of CYP1A2 Detected allele Sequence of probe 5'-3' Reference Phenotyping using caffeine showed no difference in CYP1A2*1F metabolic activity between different genotypes in nonsmokers but a significant difference in smokers homozygous for the A allele compared with the other genotypes in terms of a 1.6-fold increased activity.1-17-1 Unexpectedly, another study demonstrated a lower in vivo activity in patients with colorectal cancer than in...

Molecular Characterization Of Pathogen And Molecular Testing

E. histolytica and E. dispar genome shotgun sequencings are underway as a joint effort from TIGR 11 and the Sanger Centre, 12 funded by the National Institute for Allergy and Infectious Diseases and the Wellcome Trust, respectively. The genome is around 18-20 Mb in size in 14 chromosomes. 11-13 However, partial genomic data from these organisms are available since the end of the 1980s. 14,15 These data allowed designing molecular assays that are sensitive to detection and, more interestingly,...

Optimization Of Liposomal Delivery

Efficient in vivo nucleic acid-liposome complexes have unique features, including their morphology, mechanisms for cell and nuclear entry, targeted delivery, and ability to penetrate across tight barriers and throughout target tissues. Liposomes have different morphologies based on their composition and the formulation method that contribute to their ability to deliver nucleic acids in vivo. Formulations frequently used for the delivery of nucleic acids are lamellar structures, including small...

Management of Thalassemia Major

Patients affected by thalassemia major are treated with regular blood transfusions and iron chelation therapy with desferrioxamine B (DFO). Life expectancy with this treatment extends to the third decade. The alternative oral iron chelator deferiprone (L1) is indicated only in patients with proven allergy or toxicity from DFO. Alternative chelation strategies and drugs, including the combination of deferiprone and DFO or ICL670 alone, are under investigation. Bone marrow transplantation (BMT)...

Clinical Aspects

The incubation period for trichinellosis lasts from 2 to 50 days, depending on the number of infective larvae ingested (a greater number of larvae correspond to a shorter incubation period). 7 The acute stage of trichinellosis corresponds to the phase in which the newborn larvae migrate from the lymphatic vessels and invade the muscle cells. The acute stage can be preceded by loose stools or diarrhea, with flatulence, moderately intense abdominal pain, loss of appetite, and vomiting. 7 The...

Metabolism of Chemotherapeutic Agents

Tenable, although the failure to define precise roles for individual GST has hampered assessment of the implications of polymorphism as selection of diseases whose pathogenesis may be affected is subjective. Thus while associations between GST genotypes and clinical pheno-type have been reported they cannot be predicted, and it is unclear why some, but not other, links are significant. Further, study reproducibility is a problem, probably because of variable definition of phenotypes and...

Clinical Description Of Infection

Disease in human hosts greatly depends on the condition of the host, with more severe outcomes in immunocom-promised and malnourished individuals. 4 Most patients present profuse watery diarrhea containing mucus, 7-10 days after acquisition of infection. In immunocompetent hosts, the disease is usually self-limiting with a median duration of 9-15 days. Acute diarrhea may be resolved in a few days or may persist for 4-7 weeks. Three major presentations of symptoms are observed 1) asymptomatic...

Type Of Pathogen

Candida species are primarily saprophytic eukaryotic organisms and only occasional opportunistic pathogens. Many species exhibit a bitrophic existence, spending much of their life as common commensals of the gastrointestinal (GI) tract and mucous membranes of humans only to become pathogenic because of debilitation of the host. Indeed, the most significant source of Candida causing candidosis is endogenous. 2 Despite the main cause of invasive candidosis (IC Candida albicans) being considered...

Comparative Genomic Hybridization In Cancer Cytogenetics

Genetic alterations associated with neoplasia have been well defined in hematological malignancies by both classical and molecular cytogenetics.1-5'6-1 In contrast, there is significantly less information known about the cytogenetics and molecular cytogenetics of solid tumors. This is because of technical difficulties in the production of metaphase spreads from these tumor cells. Karyotype analysis requires viable, proliferating cells that can be arrested in the metaphase stage of the cell...

Applications in Qualitative PCR

The nPCR systems are currently widely employed in both qualitative, simple 3,4 or multiplex, 5-13 and quantitative1-14,15-1 diagnostic tests, as well as in PCR systems employed in the identification of viruses. 16-19 Regarding the above mentioned qualitative nPCRs, the primer design allows to detect from 10 to 100 genome copies per microliter of any adenovirus 4 or lyssavirus, 3 and can be performed from a variety of samples including cerebro-spinal fluid (CSF), urine, stools, respiratory...

Microsatellite Instability In Hnpcc And In Sporadic

The MSI-H phenotype is a defining feature of HNPCC patients and is a result of predisposing constitutional mutations in the MMR genes MSH2, MLH1, MSH6, hPMS2, and hMSH3. 8 Most mutations were found in MSH2 and MLH1 and less frequently in MSH6, hPMS2, and hMSH3. Germline mutations in MMR genes predispose not only to CRC but also to endometrial, upper urinary tract, and ovarian malignancies. Small intestinal and gastric cancers as well as tumors of the biliary tract are also seen in HNPCC, but...

Assay Formats Direct Hybridization

Several formats of wet-ware were adopted to bead-based nucleic acid testing. The simplest method is the direct hybridization of labeled nucleic acids to beads carrying a sequence-specific oligonucleotide. An assay for gene expression analysis using color-coded microspheres was developed using this technique. In Ref. 4 , labeled cRNA was generated which was hybridized to a set of 20 bead populations carrying specific capture sequences of 25 bp for 20 Arabidopsis genes. The obtained expression...

Impact Of Fish Analysis In Different Research Areas

More than 5 million sperm cells from more than 500 normal men have been analyzed by a number of laboratories from around the world. 7 These studies have included the frequency and distribution of aneuploidy in sperm cells, and the effect of donor age, lifestyle factors, and geographical and ethnic factors. Frequency and Distribution of Aneuploidy The frequency and distribution of disomy in human sperm cells is of interest to determine the normal baseline values and to determine if all...

Oxidative Stress Related Disorders

Oxidative stress is implicated in the inflammatory demy-elination that characterizes multiple sclerosis suggesting GST polymorphisms may be associated with disability. In 177 patients with disease duration over 10 years, GSTM3 AA (OR 2.4) and homozygosity for both GSTM1*0 and GSTP1*Ile105-encoding allele (OR 5.0) were linked with severe disability suggesting that long-term prognosis in MS is influenced by GST-mediated ability to remove toxic products of oxidative stress. 1 Exposure to...

Hcv Rna Quantitation

Quantitation of HCV RNA by bDNA technology is possible with the commercial VERSANT HCV RNA 3.0 assay. The performance characteristics of this test were established in our laboratory during the so-called ''beta trials'' before licensing and are given in Table 1. 4 Our findings were essentially confirmed by other studies, 11,12 and showed that the assay, from an analytical point of view, can be used as a routine tool for HCV RNA quantitation in clinical laboratory settings. Table 1 Analytical...

Infectious Disease Applications

The development and ongoing improvement of bDNA technology during the past decade was mainly driven by the increasing demand for quantitation of nucleic acids in clinical virology. Understanding the natural history and pathogenesis of chronic viral infections caused, for instance, by hepatitis B virus (HBV), hepatitis C virus (HCV), and human immunodeficiency virus (HIV) has been greatly supported by accurate determinations of viral load. Consequently, measurements of HBV DNA, HCV RNA, and HIV...

Molecular Genetics

The NF1 gene is located on chromosome 17 and encompasses more than 300 kb of genomic DNA with 60 exons. The protein product is referred to as ''neuro-fibromin,'' a 2818 amino acid protein. Neurofibromin includes a GTPase-activating protein (GAP) domain, which regulates the conversion of Ras-GTP to Ras-GDP. Neurofibromin therefore appears to be a regulator of Rasmediated intracellular signaling. 12 The NF1 gene behaves as a classic tumor suppressor. Affected individuals are heterozygous for NF1...

Replication And Infectivity

Orthopoxvirus genomes encode about 176 to 266 proteins, including a number of enzymes and factors that are necessary for self-replication and maturation. The central region of the genome contains highly conserved genes that are essential for viral replication, and the terminal regions contain less conserved genes that are important for virus-host interactions. Although it is well known that replication occurs in the cytoplasm, it is not clear how or if host cell nuclear factors are involved in...

Studies In Renal Disease

The individual response to treatment with ACE inhibitors is highly variable. 20 Therefore, several studies have been conducted in which the efficacy of ACE inhibitor therapy was studied in relation to ACE genotype in patients with kidney diseases (Table 2). Yoshida et al. 21 studied the response to ACE inhibitor therapy in 21 Japanese patients with IgA-nephropathy, who were treated with lisinopril (10 mg day). After 4 years of therapy, only patients with DD genotype showed a significant...

Biology Of The Organisms

Chlamydiae are obligate intracellular bacteria of limited metabolic capability, characterized by a dimorphic growth cycle. The infectious form is called the elementary body (EB) and is a condensed sporelike spheroid. The EB is metabolically inactive, consisting of a tightly compacted chromosome or nucleoid, and an outer membrane of covalently linked lipopolysaccharides. When it encounters host cells, the EB is taken in by receptor-mediated endocytosis. It has been suggested that a trimolecular...

Historical Introduction

Wilson disease, also referred to as hepatolenticular degeneration, was first described in 1912 by Samuel A. Kinnier Wilson as a familial progressive disorder of the central nervous system accompanied by liver cirrhosis, and by Bruno Fleischer as a ''pseudosclerosis'' with tremor, liver cirrhosis, and corneal deposits. 1,2 One year later, Rumpel demonstrated an excessive hepatic copper accumulation in Wilson disease. 3 This finding was confirmed by Luthy, who also detected an elevated copper...

Early Diagnosis Of Hemochromatosis Phenotyping Vs Genotyping

Early diagnosis of hemochromatosis permits treatment of iron overload and thereby prevents premature death due to hepatic cirrhosis (and primary liver cancer) and diabetes mellitus complications.1-1,2-1 Preventing iron overload may also reduce the frequency or severity of arthropathy, hypogonadotrophic hypogonadism and other endocrinopathic disorders, and cardiac abnormalities. 1,2 The cornerstone of case detection in medical care and in population screening has been the measurement of serum...

Clinical Presentation

Wilson disease is a protean disorder with a broad clinical spectrum owing to the toxic effect of copper on the various organ systems. The disorder occurs worldwide with an estimated prevalence in Caucasians of approximately 1 per 30,000 to 1 per 100,000 live births and a carrier frequency of about 1 per 90 to 1 per 150. Most patients with Wilson disease present between 5 and 35 years of age. However, Wilson disease should be included in the differential diagnosis in individuals with liver...

Other Keratin Disorders

The spectrum of keratin disorders also encompasses a number of extracutaneous diseases. Meesmann corneal dystrophy (MSD) is inherited in an autosomal-dominant fashion with incomplete penetrance. Slit lamp identification of typical fine, round cysts in the corneal epithelium aids in differentiating this disorder from other inherited corneal dystrophies. 33 Although vision is only rarely impaired to a serious degree in MSD, cyst rupture can cause corneal erosions and intermittent decrease in...

Clinical Findings

Classical branching enzyme deficiency most frequently presents during the first few months of life, with hepatosplenomegaly and failure to thrive. Progressive liver cirrhosis, with portal hypertension, ascites, esopha-geal varices, and death, usually occurs before 5 years of age. However, there are patients who have survived without apparent progressive liver disease. The neuromuscular system may also be involved. Severe cardiomyopathy as the predominant symptom has also been reported. 20 The...

Potential Future Applications

Because FISH analysis is simple and provides data on large sample sizes of sperm cells, it has been suggested as a technique for assessing the effects of potential mutagens on human sperm. We and others have studied cancer patients treated by chemotherapeutic agents and determined that these agents can cause a significant increase in the frequency of sperm chromosome abnormalities, particularly during chemotherapy and up to 2 years following treatment. 27-29 The effects of pesticides have also...

Genitourinary Chlamydial Infection

Chlamydia trachomatis infection of the genitourinary tract is the most common bacterial STD in both the United States (> 4 million new cases per year) and worldwide (> 50 million new cases per year). Many infected individuals of either sex are asymptomatic but symptomatic urethritis is common in men, whereas women may present with a mucopurulent cervicitis and acute salpin-gitis. The most serious complication of infection in men is epididymitis. Without treatment, more than 40 of infected...

Clinical Features

Patients with PWS have central muscle hypotonia that is manifested prenatally as decreased fetal movements, ab normal fetal position, and difficult delivery. The hypotonia is evident in the newborn period, accompanied by poor sucking and failure to thrive. Hypogonadism resulting in cryptorchidism and scrotal hypoplasia in males and labial hypoplasia in females are also seen in the newborn period. Hyperphagia and food-seeking behavior start at about 1 year of age and lead to obesity by 1-6...

Management Of Hbvinfected Patients

No molecular biology-based assays are necessary for the diagnosis of acute hepatitis B, which is based on serological testing. Chronic hepatitis B is defined by HBsAg persistence in serum for more than 6 months. In this setting, HBV DNA detection-quantification is necessary to determine whether or not HBV is replicating. Table 1 Commercially available HBV DNA quantification assays HBV Hybrid Capture II (Digene, Gaithersburg, MD) Versant HBV DNA (Bayer, Ludwigshafen, Germany) Cobas Amplicor HBV...

Clinical Description Dm1

Ninety percent of DM1 patients present at adulthood with delayed muscle maturation, distal muscle weakness, wasting, myotonia, cataracts, cardiac abnormalities, smooth muscle dysfunction, insulin resistance, daytime sleepiness, testicular atrophy (low reproductive fitness), ''difficult'' personality, neuropsychiatric disturbances, and frontal balding. 1 Ten percent of the patients present at infancy with hypotonia (floppy infant), oromotor dysfunction, tent-shaped mouth, feeding and respiratory...

Visceral Signs And Symptoms

Splenomegaly exists in all patients, and is typically associated with hypersplenism, early satiety, and, in children, growth (height) retardation. Anemia and throm-bocytopenia, resulting in fatigue and bleeding tendency, respectively, are among the earliest presenting signs and most prominent features of Gaucher disease. In almost all symptomatic patients, there is some degree of hepatomegaly but only rarely are liver function tests abnormal. Bone involvement is among the more variable and...

Conclusion

The discovery that subtelomeric rearrangements are a significant cause of unexplained MR has led to a vast number of studies aimed at providing diagnoses in this group. Results from over 3000 patients show that rearrangements involving the ends of chromosomes can be detected in about 5 of unselected cases, that the prevalence of subtelomeric anomalies is higher in patients with moderate to severe MR (about 7 ), and that in about half of those with subtelomeric anomalies, the rearrangements...

Method 1 Nonorganic Solvents Method Puregenetm Dna Isolation

The PUREGENE DNA Isolation kit (Gentra Systems Minneapolis, MN) is used to extract DNA from mummified tissues. 1,10 In this kit, purification of genomic DNA is carried out using ammonium acetate as a substitute for toxic organic solvents in the protein precipitation step. The pulverized tissue is mixed with cell lysis solution containing Tris-HCl, ethylenediaminetetraacetic acid (EDTA), and sodium dodecyl sulfate (SDS), and incubated at 65 C for 60 min. After proteinase K solution (20 mg mL)...

Mugpcr

Abbreviations ADPL allele discrimination by primer length PAMSA PCR amplification of multiple specific alleles CB competitive blocker CTPP confronting two-pair primer MUG multiplex universal genotyping. Abbreviations ADPL allele discrimination by primer length PAMSA PCR amplification of multiple specific alleles CB competitive blocker CTPP confronting two-pair primer MUG multiplex universal genotyping. allele-specific reaction internal control reaction Fig. 1 Principle of ARMS-PCR (see text for...

Clinical Significance Of Legionella

Legionella causes two different manifestations of pulmonary disease a mild, flulike illness, called Pontiac fever, and pneumonia. Pontiac fever has an incubation period of 1-2 days and is characterized by malaise, myalgias, fever, headache, and, sometimes, nonproductive cough. Only symptomatic therapy is required, and complete recovery within 1 week can be expected. Pneumonia, on the other hand, is the predominant manifestation of the more severe form of legionellosis, Legionnaires' disease....

The Virus

The family Astroviridae encompasses small nonenveloped viruses that infect a variety of animal species. They were originally classified among the small round structured viruses (SRSV) causing diarrhea in humans. They are round with icosahedral symmetry and 27-30 nm in diameter. They were first described by Madeley and Cosgrove 1 as a potential cause of diarrheal disease in human infants and named for the characteristic five- or six-pointed star (astron is Greek for a star), visible on the...

Multiplex Nasba And

Greijer et al. 5 designed a multiplex real-time NASBA using molecular beacon probes labeled with three different fluorophores to quantify human cytomegalovirus (CMV) IE1 mRNA by competitive coamplification of wild-type and calibrator RNA. A simultaneous detection of late pp67mRNA in whole-blood samples of CMV-infected lung transplant patients was also obtained. Despite the somewhat lower sensitivity of the real-time NASBA compared with the conventional NASBA, the simultaneous quantification of...

Retargeting Of Adenoviral Vectors

The adenovirus contains a protein capsid with protruding fibers. These fiber proteins form a trimer and the C-terminal region of these fibers is called the fiber knob. The fiber knob recognizes and binds to the host cell via the coxsackievirus-adenovirus receptor (CAR), which is present on many cell types. Upon binding to CAR, secondary interactions between the adenovirus and cell surface receptors cause the internalization of the adeno-virus. By shielding or modifying the fiber of the...

Overview Technical Description

Ramification amplification (RAM), 1,2 also termed hyperbranched rolling circle amplification1-3-1 or cascade rolling circle amplification,1-4-1 utilizes a closed C-probe or padlock probe and a DNA polymerase under isothermal conditions. The choice of DNA polymerase is based on the following requirements a good strand displacement activity in order to separate downstream DNA from the template strand and a high processivity (i.e., incorporating nucleotides continuously on a given primer without...

Tobacco Related Diseases

GSTM1 and GSTT1 have been particularly studied as risk candidates for tobacco-related cancers. 14,15 While some studies show significantly increased risk for GSTM1*0 and GSTT1*0 homozygotes, others have not replicated these findings prompting the use of meta-analysis. For example, analysis using 43 studies indicates that GSTM1 null is not associated with increased lung cancer risk and that there is no evidence for an interactive effect between the genotype and tobacco consumption.1-15-1...

Nat And Disease

The role of NATs in human disease has recently been extensively reviewed. 3 The following is a summation of findings to date. The association between acetylator status and the risk of various diseases has been extensively reported. Altered risk with either the slow or rapid phenotype has been observed for bladder, colon and breast cancer, systemic lupus erythematosis, diabetes, age-related cataracts, Gilbert's disease, Parkinson's disease, and Alzheimer's disease. These associations imply a...

Practical RCA formats

Generally, the RCA-based diagnostics can be classified into two groups some of them operate with the preformed circular probes Fig. 1D and E for the peptide nucleic acid (PNA)-assisted, nick-induced RCA and immuno-RCA, Fig. 1 Schematics of different RCA processes the arrowhead symbolizes the DNA polymerase. Given a small, < 100-nt size of DNA minicircles used in RCA and the strong rigidity of dsDNA fragments with these lengths, only part of the circular probe can be base-paired at any given...

Pathophysiologic And Genetic Features

The first step in the telangiectasia formation consists a focal dilatation of postcapillary venules with predominant stress fibers in pericytes along the luminal border. As the venules enlarge, they become convoluted and form excessive layers of smooth muscle cells without elastic fibers and connect directly with the dilated arterioles.1-1-1 The molecular basis consists of a defect of the TGF-p signaling pathway which, in normal conditions, plays an important role in vascular remodeling and...

Conclusion Genomics And Vaccination

Despite the advances associated with molecular techniques, meningococcal disease remains a globally significant health problem, and the prospect of comprehensive vaccination remains elusive. Whereas vaccines based on the capsular polysaccharide are either licensed or under development against meningococci associated with serogroups A, C, and Y, as well as W-135, attempts to develop a polysaccharide-based serogroup B vaccine have proven unsuccessful. This fact is especially relevant as much of...

Principles Of Rna Storage

Basically, RNA can be stored in purified form after its extraction or synthesis, or unprocessed samples containing the RNA of interest are stored. However, even in the latter case, proper care must be taken to avoid (re) contamination of RNA with RNases once it is extracted from the stored sample. Little information is available on the long-term stability of isolated and purified RNA during storage. Most studies have been carried out focusing on various storage conditions of unprocessed samples...

Principle Of The Taqman Assay

Like several eubacterial DNA polymerases, Thermus aquaticus (Taq) polymerase is a bifunctional enzyme consisting of a polymerization domain and a nuclease domain, and it is the latter that is crucial for the TaqMan assay. 1 In essence, the TaqMan assay is identical to a standard PCR reaction powered by Taq polymerase, except for the addition of the TaqMan probe, a nonextendable fluorescently labeled oligonucleotide hy bridizing on the target DNA between the forward and reverse primers 2-3 (Fig....

Sample Preparation for PFGE

Standard sample preparation procedures for DNA isolation involving mechanical forces are inappropriate for the analysis of fragile, high-molecular weight DNA molecules that easily break into small pieces. To prevent DNA damage, intact cells mixed with warmed, liquid agarose are pipetted into plug molds, 10 x 5 x 1.5 mm in size. Cells embedded in the agarose plugs are lysed in situ by detergents and enzymes. The agarose matrix keeps the large DNA molecules intact, while permitting diffusion of...

Kras

Associated malignancies and their penetrance Associated malignancies and their penetrance Gastric cancer (70 ) Lobular breast cancer in females (39 ) Colorectal cancer (70-85 ) Endometrial cancer (40-60 ) Ovarian cancer (12 ) Stomach, uroepithelial, hepatobiliary, brain, small bowel cancers Colorectal cancer Ovarian stromal tumors Uterine and cervical cancer Gastrointestinal tract polyposis and cancers, breast and testis, cancers Thyroid cancer Breast cancer Gastric cancer Leukemia Breast...