Locally advanced colorectal cancer in pregnancy.

OBJECTIVE: To highlight the issue of diagnostics and treatment management in colorectal cancer of pregnant women. DESIGN: Case report. SETTING: Department of Obstetrics and Gynaecology, Pardubice Hospital. CASE REPORT: We present a case of a 24-year-old secundigravida, diagnosed as colorectal cancer in week 34 of pregnancy, manifesting as a diarrhoea and a stomach ache. The diagnosis was made in an advanced stage due to underestimation of clinical symptoms. CONCLUSION: The diagnosis of the colorectal cancer in pregnancy can be difficult because the presenting symptoms are easily attributable to pregnancy. Therefore the advanced disease is diagnosed more frequently in pregnant patients than in the general population. Colorectal cancer should be managed by multidisciplinary team with regard to gestation.

[Spontaneous rupture of the uterine artery in connection with pregnancy]. / Spontánní ruptura uterinní arterie v souvislosti s tehotenstvím.

OBJECTIVE: To present two cases of spontaneous uterine artery rupture in connection with pregnancy, accompanied by life endangering bleeding. In the first case we talk about a thirty-three years old primigravida nullipara in the 20th week of pregnancy. In the second case there is a woman, who gave spontaneous birth in the 36th week of pregnancy. DESIGN: Case report. SETTING: Department of Obstetrics and Gynecology, Pardubice Hospital. METHODS: Own observation. CONCLUSION: Spontaneous rupture of the uterine artery in connection with pregnancy is a rare complication that can be fatal for mother and fetus. Clinical symptoms are often nonspecific, diagnostics is difficult, and diagnosis is often made during the surgery. It is necessary to include this rare complication in the differential diagnosis in pregnant women with abdominal pain and haemoperitoneum of unknown etiology and in postpartal women with an unknown source of bleeding.

Primary malignant melanoma of the vagina, case report and review of the literature.

OBJECTIVE: Presentation of primary malignant melanoma of the vagina. Summary of clinical findings and management. DESIGN: Case report. SETTING: Nemocnice Pardubického kraje a.s., Porodnicko-gynekologická klinika, Pardubice. OBSERVATION: We report a case of primary malignat melanoma in a 44 year old woman, with symptoms of abnormal vaginal bleeding and painful intercourse. CONCLUSION: Primary malignant melanoma of the vagina is rare disease. Goal of therapy is complete removal of primary tumor.

[Suburethral sling removal due to chronic pain]. / Exstirpace suburetrální pásky z duvodu chronických bolestí.

OBJECTIVE: To point out one possible complication after suburethral tape insertion. To present methods of treatment, especially partial or total removal of the tape. Few successful cases of treatment are shown in case reports. DESIGN: Case report. SETTING: Department of obstetrics and gynecology, Pardubice hospital. OBSERVATION: Chronic pain after insertion of suburethral tape is a rare complication. There are several possible causes, such as tape erosion with chronic inflamation, post-colposuspension syndrome, myo-fascial syndrome, nerve damage during tape implantation or due to overproduction of fibrous tissue. Partial or total tape removal is needed when the conservative treatment fails. Patients may remain continent or a new tape is placed. Pain relief is usually complete but there are times when additional treatment is needed. CONCLUSION: Sling removal is a successful method of treating chronic pain after suburethral tape insertion. Patients describe plain relief and improvement of quality of life.

Golden SusPtrit: a genetically well transformable barley line for studies on the resistance to rust fungi.

KEY MESSAGE: We developed 'Golden SusPtrit', i.e., a barley line combining SusPtrit's high susceptibility to non-adapted rust fungi with the high amenability of Golden Promise for transformation. Nonhost and partial resistance to Puccinia rust fungi in barley are polygenically inherited. These types of resistance are principally prehaustorial, show high diversity between accessions of the plant species and are genetically associated. To study nonhost and partial resistance, as well as their association, candidate gene(s) for resistance must be cloned and tested in susceptible material where SusPtrit would be the line of choice. Unfortunately, SusPtrit is not amenable to Agrobacterium-mediated transformation. Therefore, a doubled haploid (DH) mapping population (n = 122) was created by crossing SusPtrit with Golden Promise to develop a 'Golden SusPtrit', i.e., a barley line combining SusPtrit's high susceptibility to non-adapted rust fungi with the high amenability of Golden Promise for transformation. We identified nine genomic regions occupied by resistance quantitative trait loci (QTLs) against four non-adapted rust fungi and P. hordei isolate 1.2.1 (Ph.1.2.1). Four DHs were selected for an Agrobacterium-mediated transformation efficiency test. They were among the 12 DH lines most susceptible to the tested non-adapted rust fungi. The most efficiently transformed DH line was SG062N (11-17 transformants per 100 immature embryos). The level of non-adapted rust infection on SG062N is either similar to or higher than the level of infection on SusPtrit. Against Ph.1.2.1, the latency period conferred by SG062N is as short as that conferred by SusPtrit. SG062N, designated 'Golden SusPtrit', will be a valuable experimental line that could replace SusPtrit in nonhost and partial resistance studies, especially for stable transformation using candidate genes that may be involved in rust-resistance mechanisms.

Time-lapse imaging of the initiation of pollen embryogenesis in barley (Hordeum vulgare L.).

Pollen embryogenesis provides exciting opportunities in the areas of breeding and biotechnology as well as representing a convenient model for studying the process of plant cell proliferation in general and embryogenesis in particular. A cell culture system was devised in which immature barley pollen could be cultured as a monolayer trapped between the bottom glass-cover slip of a live-cell chamber and a diaphanous PTFE membrane within a liquid medium over a period of up to 28 d, allowing the process of embryogenesis to be tracked in individual pollen. Z-stacks of images were automatically captured every 3min, starting from the unicellular pollen stage up to the development of multicellular, embryogenic structures. The method should prove useful for the elucidation of ultrastructural features and molecular processes associated with pollen embryogenesis.

Convergent evolution of a metabolic switch between aphid and caterpillar resistance in cereals.

Tailoring defense responses to different attackers is important for plant performance. Plants can use secondary metabolites with dual functions in resistance and defense signaling to mount herbivore-specific responses. To date, the specificity and evolution of this mechanism are unclear. Here, we studied the functional architecture, specificity, and genetic basis of defense regulation by benzoxazinoids in cereals. We document that DIMBOA-Glc induces callose as an aphid resistance factor in wheat. O-methylation of DIMBOA-Glc to HDMBOA-Glc increases plant resistance to caterpillars but reduces callose inducibility and resistance to aphids. DIMBOA-Glc induces callose in wheat and maize, but not in Arabidopsis, while the glucosinolate 4MO-I3M does the opposite. We identify a wheat O-methyltransferase (TaBX10) that is induced by caterpillar feeding and converts DIMBOA-Glc to HDMBOA-Glc in vitro. While the core pathway of benzoxazinoid biosynthesis is conserved between wheat and maize, the wheat genome does not contain close homologs of the maize DIMBOA-Glc O-methyltransferase genes, and TaBx10 is only distantly related. Thus, the functional architecture of herbivore-specific defense regulation is similar in maize and wheat, but the regulating biosynthetic genes likely evolved separately. This study shows how two different cereal species independently achieved herbivore-specific defense activation by regulating secondary metabolite production.

Tumor necrosis factor and lymphotoxin gene expression in human tumor cell lines.

In this paper we show that eight of 17 tumor cell lines of various tissue origin constitutively express tumor necrosis factor (TNF) mRNA. Five of these eight cell lines concomitantly contained lymphotoxin (LT) mRNA. Of the remaining nine cell lines that lacked detectable TNF or LT gene expression, five could be induced by phorbol ester and/or cytokines to accumulate the respective mRNAs. While TNF mRNA was found not only in neoplastic hematopoietic cells, but also in cell lines derived from carcinomas, LT gene expression seemed to be restricted to lymphoid tumor cells. Tumor cells that expressed LT mRNA also secreted LT protein and proved to be resistant to the cytostatic/cytotoxic effects of their own protein product as well as to exogenous recombinant TNF and recombinant LT. In contrast, most of the cell lines containing TNF mRNA did not release TNF protein into the supernatants, indicating that TNF gene expression may be controlled predominantly at a post-transcriptional level. Thus, the presence of TNF mRNA does not necessarily reflect a TNF-resistant phenotype. Our findings demonstrate that TNF and/or LT mRNA expression is a rather common phenomenon in long-term cultured tumor cell lines and reveal that ectopic TNF and/or LT production by tumor cells may be involved in certain paraneoplastic syndromes as well as in tumorigenesis.

Characterization of an Krox-24/Egr-1-responsive element in the human tumor necrosis factor promoter.

We have analyzed in various human leukemic cell lines a previously unrecognized region within the human TNF gene promoter that contains the sequence motif 5'-CCGCCCCCGCG-3'. This GC-rich sequence maps to bps -170 and -160 of the TNF gene. Electrophoretic mobility shift assays (EMSA) combined with methylation interference analysis revealed the binding of two distinct proteins with overlapping recognition sites. Supershift assays identified the constitutive transcription factor Sp1 and the immediate-early growth-response transcription factor Egr-1/Krox-24. Interestingly, this Egr-1-related factor was induced by PMA but not by TNF. The TNF gene GC-rich sequence conferred PMA responsiveness when linked to a heterologous minimal c-fos promoter. To examine the involvement of Egr-1/Krox-24 in TNF gene regulation, a Krox-24 expression vector was used, pSCTKr24. In Jurkat T cells pSCTKr24 stimulated pTNF-286CAT that contains sequences -286 to +34 of the human TNF gene fused to the chloramphenicol acetyltransferase (CAT) gene. Moreover, pSCTKr24 also stimulated the TNF gene GC-rich sequence linked to the minimal c-fos promoter. However, deletion of this site did not result in markedly reduced TNF promoter activity, suggesting that the Egr-1/Krox-24 response element may play an auxiliary role in TNF gene regulation.

The green fluorescent protein targets secretory proteins to the yeast vacuole

The green fluorescent protein (GFP) was used as a marker to study the intracellular transport of vacuolar and secretory proteins in yeast. Therefore, the following gene constructs were expressed in Saccharomyces cerevisiae under control of the GAL1 promoter: GFP N-terminally fused to the yeast secretory invertase (INV-GFP), the plant vacuolar chitinase (CHN-GFP) and its secretory derivative (CHNDeltaVTP-GFP), which did not contain the vacuolar targeting peptide (VTP), both chitinase forms (CHN and CHNDeltaVTP), GFP without any targeting information and two secretory GFP variants with and without the VTP of chitinase (N-GFP-V and N-GFP). Whereas chitinase without VTP is accumulated in the culture medium the other gene products are retained inside the cell up to 48 h of induction. Independently of a known VTP they are transported to the vacuole, so far as they contain a signal peptide for entering the endoplasmic reticulum. This was demonstrated by confocal laser scanning microscopy, immunocytochemical analysis and subcellular fractionation experiments as well. The transport of the GFP fusion proteins is temporary delayed by a transient accumulation in electron-dense structures very likely derived from the ER, because they also contain the ER chaperone Kar2p/Bip. Our results demonstrate that GFP directs secretory proteins without VTP to the yeast vacuole, possibly by the recognition of an unknown vacuolar signal and demonstrates, therefore, a first limitation for the application of GFP as a marker for the secretory pathway in yeast.

Hormonal regulation of protein disulfide isomerase and chaperone synthesis in the rat exocrine pancreas.

Folding and assembly of polypeptides translocated into the rough endoplasmic reticulum (RER) is facilitated by a set of resident proteins in the lumen of the RER. We studied the regulation of synthesis of the RER luminal proteins immunoglobulin heavy chain binding protein (BiP) and protein disulfide isomerase (PDI), and of the cytosolic stress 70 protein (hsc70) after hormonal stimulation of the pancreatic exocrine secretory pathway. Their rate of synthesis was assessed at both mRNA and protein levels and under two experimental conditions that are associated with large increases in exocrine production. After in vivo stimulation of the pancreas by either endogenous release of cholecystokinin (CCK) following proteinase inhibitor feeding (FOY-305) or by in vivo infusion of the pancreatic secretagogue cerulein, the relative rates of synthesis detected for BiP and PDI were enhanced 2.5 to 4-fold compared to control. Interestingly, the kinetics and the degree of hsc70 mRNA induction were almost identical to those of BiP and PDI, suggesting coordinated hormonal regulation of BiP, PDI as hormonal stimulation was even twice that following heat shock treatment. The mRNA levels of calreticulin (CaBP3) increased up to 2.3-fold with a kinetic comparable to that of BiP, PDI and hsc 70, while CaBP1 and the RER membrane proteins, ribophorin I and the signal recognition particle receptor did not show any changes in their relative mRNA amounts after hormonal stimulation. The increase in the rates of PDI and chaperone biosynthesis exceeds the associated increase in total protein biosynthesis. In vitro experiments, using transformed rat acinar cells (AR4-2J) in which pancreatic enzyme synthesis can be induced by glycocorticoid hormones, demonstrated that induction of PDI and chaperone mRNA synthesis preceded extensive mRNA expression of secretory proteins.

Regulated secretion of mature cathepsin B from rat exocrine pancreatic cells.

By indirect immunofluorescence and immunogold electron microscopy with an antibody that recognizes specifically the two forms of native mature rat cathepsin B (31 kDa and 5:25 kDa) but not the proenzyme, we detected cathepsin B not only in lysosomes of adult rat exocrine pancreatic cells but also in the trans Golgi condensing vacuoles, the zymogen granules and the pancreatic juice in the intralobular ducts. In contrast, immunocytochemistry with an antibody specific for rat cathepsin D showed the latter to be present in the same cells only in lysosomal compartments as expected. The same pattern of labeling with these two antibodies was found in the first zymogen granules to form in 17-day-old fetal rat pancreas. Counts of the extent of immunogold labeling of cathepsin B in the adult exocrine cells showed that the concentration of the enzyme was only two-fold higher in the lysosomal compartments than in the zymogen granules. To confirm these observations, rat pancreatic postnuclear supernatant (PNS), a fraction enriched in zymogen granules and rat pancreatic juice obtained by catheterization of the pancreatic duct, were subjected to 2D gel electrophoresis followed by immunoblotting with the cathepsin B antibody. All three samples contained a 31 kDa protein recognized by the antibody with a pI of about 4.5, the single chain mature form of cathepsin B. We then radiolabeled pancreatic PNS and zymogen granule fractions with benzyloxycarbonyl-Tyr[125I]-Ala-CHN2, an affinity label that covalently binds to the active sites of mature forms of both cathepsin B and cathepsin L. In both PNS and zymogen granule fractions this reagent labeled cathepsin B. Immunoprecipitation experiments showed that the antibody to cathepsin B recognized specifically both the single chain and the double chain mature forms of cathepsin B in the native state. Finally, Northern blots with a cDNA of rat cathepsin B showed that the concentration of cathepsin B mRNA in total pancreatic RNA increased following in vivo stimulation of the exocrine pancreatic cells with optimal doses of cerulein, a cholecystokinin analogue. We conclude that significant amounts of mature cathepsin B are secreted from exocrine pancreatic cells via the apical regulated exocytotic pathway, and we discuss this in terms of models for sorting of proteins to the cores of dense cored secretory granules.

The Effects of Suburethral Tape on the Symptoms of Overactive Bladder.

Suburethral tension-free vaginal tape is used for the treatment of stress urinary incontinence with a high success rate. Often patients report having stress incontinence, as well as co-existing micturition problems which are attributable to overactive bladder syndrome (OAB). The present study examines the effect of suburethral tape on the symptoms of OAB. In the study, we used the transobturator vaginal tape inside-out technique (TVT-O). Materials and Methods: 53 patients were included in the study, all had proven urodynamic stress incontinence and symptoms of overactive bladder. The patients were examined preoperatively and 3 months after the TVT-O placement. Results: The individual OAB symptoms improved significantly, with urinary frequency and urge incontinence improving more than nocturia. The frequency of micturition decreased on average from 16.1 to 10.1 episodes/24 hours, while nocturnal frequency of micturition decreased from 2.2 to 1.1. Not a single patient experienced the simultaneous worsening of all three measured variables, however 19 % of patients did report their simultaneous disappearance. Their quality of life that had been affected by OAB was measured on the basis of validated questionnaires, and found to have improved significantly. Only 28 % of patients reported a desire for drug treatment of OAB symptoms following tape placement. Conclusions: TVT-O placement leads to a significant improvement of the symptoms of overactive bladder syndrome. Patient quality of life - which was affected by OAB - was also enhanced by the tape placement. This accounts for a substantial share of the overall success of the suburethral tape.

O-Acetylserine sulfhydrylase and S-sulfocysteine synthase activities of Rhodospirillum tenue.

O-Acetylserine sulfhydrylase in cell-free extracts of Rhodospirillum tenue was markedly repressed after growth in the presence of sulfide or thiosulfate, whereas S-sulfocysteine synthase activity remained almost unchanged. Purification on DE52 cellulose resulted in the separation of two proteins: Protein I with a molecular weight of 57000 had O-acetylserine sulfhydrylase activity only, while protein II with a molecular weight of 46000 had S-sulfocysteine synthase activity in addition. The activity of protein II with O-acetylserine plus sulfide was about 1.5 of that with O-acetylserine plus thiosulfate. Protein I from sulfate-grown cells possessed 74% of the total O-acetylserine sulfhydrylase, protein II 26%. Growth with sulfide repressed only the synthesis of protein I, which after separation showed only 19% of the measurable O-acetylserine sulfhydrylase, whereas protein II now possessed 81%. Regulatory and kinetic phenomena of the two activities were studied. In addition to the phototrophic bacteria studied earlier, also Rhodomicrobium vannielii, Rhodopseudomonas acidophila, Rhodocyclus purpureus and Thiocystis violacea were found to contain O-acetylserine sulfhydrylase activities; the latter two species contained S-sulfocysteine synthase activities in addition.