Genomic structure of proton-coupled oligopeptide transporter hPEPT1 and pH-sensing regulatory splice variant

Proton-coupled oligopeptide transporter PEPT1 facilitates the transport of dipeptides and peptoid drugs (including antibiotics) across the cell membranes of endothelial and epithelial cells. Substrate transport by the proton symport is driven by pH gradients, while the profile of pH sensitivity is regulated by a closely related protein. hPEPT1-RF. We investigated the genomic structure of hPEPT1 and hPEPT1-RF. Analysis of the high-throughput genomic sequence (HTGS) database revealed that hPEPT1 and hPEPT1-RF are splice variants encoded by the same gene located in chromosome 13, consisting of 24 exons. hPEPT1 is encoded by 23 exons and hPEPT1-RF by 6 exons. Coding sequences of hPEPT1-RF share 3 exons completely and 2 exons partially with hPEPT1. The genomic organization of hPEPT1 shows high similarity with its mouse orthologue. Exon-intron boundaries occur mostly in the loops connecting transmembrane segments (TMSs), suggesting a modular gene structure reflecting the TMS-loop repeat units in hPEPT1. The putative promoter region of hPEPT1 contains TATA boxes and GC-rich regions and a potential insulin responsive element.     

A new triphenylethylene compound,Fc-1157a

Summary The antitumor effects of a new antiestrogen, Fc-1157a¹ have been studied in vitro and in vivo. In vitro the effect of Fc-1157a was comparable to that of tamoxifen. The effect was dose-dependent, and at concentrations higher than 10^-6 mol/l Fc-1157a induced real cell death of the MCF-7 cells. In DMBA-induced mammary cancer in rats Fc-1157a decreased the number of new tumors and inhibited the growth of existing tumors, these effects being statistically highly significant. The ratio of growing tumors to stable and regressing tumors was significantly decreased. Although these effects were slightly stronger with Fc-1157a than with tamoxifen, the difference between these two compounds was not statistically significant.Murine uterine sarcoma, an estrogen receptor-negative tumor, was resistant to tamoxifen, but was statistically significantly inhibited by high doses (100 and 200 mg/kg^-1 day^-1 for 5 days) of Fc-1157a.The antitumor effects of Fc-1157a are due mainly to the antiestrogenic activity. At high concentrations in vitro and at high doses in vivo Fc-1157a exerts antitumor effects some of which are different from those of tamoxifen and are directed even against estrogen receptor-negative tumors. The exact mechanism of the observed cytolytic effect at high doses is unknown.     

Different synergistic roles of small polyethylenimine and Dosper in gene delivery.

Low-molecular-weight PEIs and cationic liposomes can be combined resulting in a synergistic increase in transfection efficiency as we have reported earlier. Here, we have further investigated the potential mechanisms of this synergy. Complex morphology, complex sizes and DNA condensation were studied using transmission electron microscopy, light scattering methods and ethidium bromide exclusion, respectively. Cellular uptake, transfection efficiency, and effect of proton pump inhibitor bafilomycin A1 were examined in cell cultures. The cellular uptake of DNA was negligible with PEI2K-DNA complexes, whereas the uptake of the PEI2K-DNA-Dosper or the Dosper-DNA complexes was maximally about 40%. The number of transfected cells was two times higher with PEI2K-DNA-Dosper complexes than with Dosper-DNA complexes. The PEI2K-DNA-Dosper combination was slightly less sensitive to bafilomycin A1 than the PEI25K-DNA or Dosper-DNA complexes. There were no differences between PEI2K and PEI25K in DNA condensation. Dosper condensed DNA slightly more in PEI2K complexes. The PEI25K-DNA complexes were much smaller (<250 nm) than the PEI2K-DNA complexes (0.5-12 micro m) which were also rather polydisperse. It is suggested that two independent mechanisms would lead to synergistic transfection efficiency: (1) Dosper improves the cellular uptake of PEI2K-DNA complexes, and (2) PEI2K improves a transfer of the complexes from lysosomes to nucleus.     

Extracellular and intracellular factors influencing gene transfection mediated by 1,4-dihydropyridine amphiphiles

Double-charged 1,4-dihydropyridine (1,4-DHP) amphiphiles have been shown to condense DNA and efficiently transfect it into cells in vitro [Hyvönen et al., Biochim. Biophys. Acta 1509 (2000) 451]. Alkyl chain length and buffering capacity at endosomal pH range (5.0–7.4) affected complexation and transfection activity. In this study we examined how those chemical modifications of amphiphile–DNA complexes (amphiplexes) affect their interactions with extracellular polyanions (glycosaminoglycans, albumin) and lipid bilayers, their cellular uptake and intracellular distribution. To evaluate cellular uptake, CV1-P cells were incubated with labeled DNA–amphiphile complexes and analyzed by flow cytometry. Confocal laser fluorescence microscopy was used to investigate the intracellular distribution of amphiplexes. The results showed that biophysical properties of compounds can be changed by slight structural modifications. These factors determine the intracellular kinetics and transfection efficacy of the compounds. Some extracellular glycosaminoglycans and serum interfere with 1,4-DHP-amphiphile-mediated transfection by destabilizing the amphiplexes. Neither high cellular uptake, membrane destabilizing activity nor buffering capacity alone is adequate for high transfection efficacy. The activity results from complex interplay of various factors that determine intracellular kinetics and, consequently, transfection.     

Lumbar spinal stenosis patients are satisfied with short-term results of surgery - younger age, symptom severity, disability and depression decrease satisfaction

Purpose. To study the lumbar spinal stenosis (LSS) patients' (n = 98) satisfaction with surgery outcome and associated factors at three months post-operative stage.

Method. LSS-related physical functioning and pain were assessed with Oswestry disability index, Stucki questionnaire, Visual Analogue Scale and pain drawings. Depression was assessed with 21-item Beck Depression Inventory. Psychological well-being was assessed with the Life satisfaction scale, Toronto Alexithymia Scale and Sense of Coherence Scale. All questionnaires were administered before and 3 months after surgical treatment of LSS. Satisfaction with surgery outcome was assessed with a separate scale.

Results. Considerable improvement was evident in all the functional and pain-related variables. Two-thirds (66%) of the patients were at least clearly satisfied with the surgery outcome. Younger age, symptom severity, disability and depression were independently associated with dissatisfaction with surgery outcome.

Conclusion. The lack of physical, functional and emotional well-being is associated with the patients' dissatisfaction with the surgery outcome. Patient satisfaction is a valid outcome to be measured in LSS patients undergoing surgery. It is recommended that patients should be provided with realistic pre-operative patient information and that depression be assessed pre-operatively.     

Large-scale commodity agriculture exacerbates the climatic impacts of Amazonian deforestation

In the Amazon rainforest, land use following deforestation is diverse and dynamic. Mounting evidence indicates that the climatic impacts of forest loss can also vary considerably, depending on specific features of the affected areas. The size of the deforested patches, for instance, was shown to modulate the characteristics of local climatic impacts. Nonetheless, the influence of different types of land use and management strategies on the magnitude of local climatic changes remains uncertain. Here, we evaluated the impacts of large-scale commodity farming and rural settlements on surface temperature, rainfall patterns, and energy fluxes. Our results reveal that changes in land–atmosphere coupling are induced not only by deforestation size but also, by land use type and management patterns inside the deforested areas. We provide evidence that, in comparison with rural settlements, deforestation caused by large-scale commodity agriculture is more likely to reduce convective rainfall and increase land surface temperature. We demonstrate that these differences are mainly caused by a more intensive management of the land, resulting in significantly lower vegetation cover throughout the year, which reduces latent heat flux. Our findings indicate an urgent need for alternative agricultural practices, as well as forest restoration, for maintaining ecosystem processes and mitigating change in the local climates across the Amazon basin.

During the past 50 y, ∼20% of the Amazon forest has been lost to deforestation (1, 2). These changes in the land surface have affected the functioning of ecosystems and the climate in ways we are only starting to understand. Deforestation size, for instance, is a potential factor defining the magnitude and characteristics of changes in local climate associated with forest loss (3, 4). There is also evidence that the different land uses that follow deforestation can regulate the magnitude of changes in surface energy balance and water cycle (5). Historically, there has been large variation in the characteristics and causes of deforestation (1, 6–9). In the area known as the “arc of deforestation,” two major processes have contributed to forest loss: government-supported rural settlements and expansion of market-focused large-scale agriculture (hereinafter referred to as “commodity agriculture”) (10, 11). Deforestation caused by these two types of farming systems has distinct characteristics, and each can have several variants.Rural settlements are generally associated with government colonization projects, migratory flow incentives, and the construction of new roads (7). In areas dominated by rural settlements, small properties with plots ranging from 25 to 100 ha are predominant (8, 9, 12). However, medium-sized properties ranging from 250 to 1,000 ha and farms larger than 1,000 ha may also occur. Activities inside these areas are characterized by livestock production (extensive pastures), small-scale crop production, and family farming (13). The establishment of small farms along main highways and secondary roads results in the well-known “fish bone” deforestation pattern.Forest areas taken by large-scale commodity agriculture represent a more recent stage of occupation, usually associated with spontaneous and economical migration but also, with changes in land use policies and market conditions (14). Agricultural activities aimed at commodity crop plantation are in general productive and often technologically advanced. The most common commodity crops in the Amazon region are soybean, maize, sorghum, and cotton. Nonetheless, forests are typically not converted directly into croplands, with pastures often used as a transitory land use. Permanent mid- to large-scale cattle ranching also occurs, although many of these areas are being rapidly converted into croplands (6, 14–16). Farm sizes can reach several thousand hectares. Properties are, therefore, bigger and more isolated, in comparison with rural settlements (13).Given the different characteristics of commodity agriculture and rural settlements, the spatiotemporal patterns of land cover biophysical properties can also differ considerably. In general, commodity crops cultivation involves an intensive use of the land, sometimes with two or more harvests per year (17). Hence, rapid changes in the vegetation cover, albedo, and evapotranspiration (ET) can occur (5, 18). On the other hand, in areas where small-scale pastures and agriculture are prevalent, the biophysical properties of the land surface are expected to vary less, given the less intensive use of the land (e.g., associated with family farming and agroforestry). Furthermore, modeling studies suggest that the type of vegetation involved in land cover conversions is important in determining the sign of the land change impacts (19). However, empirical studies are crucially needed to better understand how different land uses across the Amazon region affect the local and regional climate.Tropical deforestation has deep impacts on biophysical processes (1, 20–22), contributing to amplifying diurnal temperature variability (1.95 ± 0.08 °C) as well as increasing mean air temperature (∼1 °C) (23). The causes of increase in temperature are dominated by nonradiative mechanisms, in particular a decrease in latent heat flux (LE) (24). The cooling effects of albedo increase due to deforestation are in most cases outweighed by the warming effects of decreasing ET, leading to net warming (23–25).The impacts of Amazon deforestation on rainfall patterns are not yet fully understood (4). In the initial phases of deforestation, vegetation loss was shown to increase regional cloudiness and precipitation (3). In comparison with deforested areas, the greater humidity over forests leads to more convective available potential energy, which makes the atmospheric boundary layer more unstable (26). Conversely, small deforestation patches showed more active shallow convection, explaining the higher frequency of shallow clouds over deforested areas (26). However, it is unclear how these mechanisms change as deforested areas increase and land cover becomes more uniform. One hypothesis is that convective lifting mechanisms will lose force, and shallow clouds over deforested areas will no longer be favored. Modeling studies indicate that this shift is already happening in some parts of the Amazon, where deforestation has reached a point in which thermally dominated regime has declined, leading to a more dynamically driven hydroclimatic regime (27). A dynamically driven regime becomes dominant when differences in surface roughness between forest and forest clearings start to play a larger role in the atmospheric response, in comparison with the differences in the surface energy partitioning (28).As observational and modeling studies indicate that land use and management can play an important role in the climate system, overlooking these landscape heterogeneities can hinder an adequate response to the threats posed by human activities (29). Clarifying the climatic impacts of different land uses in the Amazon is crucial to foster informed plans for sustainable land management, in particular those aiming at strategies for climate change mitigation, maintenance of ecological functioning, and guarantying provision of essential ecosystem services. Here, we hypothesize that forest conversion to large-scale commodity agriculture is more detrimental to local climate than conversion to rural settlements. To test this hypothesis, we first evaluated whether or not land uses associated with commodity agriculture and rural settlements lead to quantitatively distinguishable land cover spatiotemporal patterns in regions with similar deforestation rates (1985 to 2018) and total deforested area in 2018. Next, we collected empirical evidence on how forest clearing associated with these two causes has affected local rainfall, surface temperature, and LE.     

The clinical utility of serum anti‐Müllerian hormone in the follow‐up of ovarian adult‐type granulosa cell tumors—A comparative study with inhibin B

Ovarian adult‐type granulosa cell tumors (AGCTs) require prolonged follow‐up, but evidence regarding the optimal follow‐up marker is lacking. The objective of our study was to validate the clinical usefulness of serum anti‐Müllerian hormone (AMH) and the current marker inhibin B as single and combined markers of AGCTs. We conducted a longitudinal, partially prospective cohort study of 123 premenopausal and postmenopausal AGCT patients with a median follow‐up time of 10.5 years (range 0.3–50.0 years). Serum AMH and inhibin B levels were measured from 560 pretreatment and follow‐up serum samples by using immunoenzymometric assays. We found that serum AMH and inhibin B levels were significantly elevated in patients with primary or recurrent AGCTs. The levels of both markers positively correlated to tumor size (p < 0.05). AMH and inhibin B performed similarly in receiving operator characteristic analyses; area under the curve (AUC) values were 0.92 [95% confidence interval (CI) 0.88–0.95] for AMH, and 0.94 (95% CI 0.90–0.96) for inhibin B. AMH was highly sensitive (92%) and specific (81%) in detecting a macroscopic AGCT. However, in AUC comparison analyses, the combination of the markers was superior to inhibin B alone. In conclusion, serum AMH is a sensitive and specific marker of AGCT, and either AMH or inhibin B can be monitored during follow‐up. However, combining AMH and inhibin B in AGCT patient follow‐up improves the detection of recurrent disease.     

Feasibility of abdominal plain film images in evaluation suspected drug smuggler

Objective

Drug smuggling in the gastrointestinal tract has soared within the last 20 years. Though illegal substances in the gastrointestinal tract can be visualized with ultrasound, MRI and CT, the abdominal radiograph has by far remained the most frequently used way of detecting smuggled drugs. The purpose of the study was to evaluate the inter-radiologist interpretation error and the reliability of the abdominal radiograph in detecting smuggled drugs.

Materials and methods

A total of 279 abdominal radiographs of suspected smugglers were classified by three radiologists as clearly positive or negative for drug smuggling. All available information about the cases was collected from the customs officers and police.

Results

Out of these cases 203 (73%) were interpreted as negative and 35 (13%) as positive by all three radiologists. In 86% of the cases there was, therefore, an inter-radiological agreement in interpreting the images. In 41 (14%) cases, however, there was an inter-radiologist disagreement. Kappa-value for inter-observer variability was 0.70.

Conclusions

In up to a seventh of the abdominal radiographs the interpretation can be challenging even for an experienced radiologist. False positive interpretation can lead to innocent passengers being detained in vain. As negatively interpreted images usually result in releasing of the suspect, there is no way of knowing how many false negative occur. This makes the abdominal radiograph a suboptimal examination, and low dose CT should be considered as the screening modality for gastrointestinal drug smugglers.     

Erratum to: Long-term results of surgery for lumbar spinal stenosis: a randomised controlled trial

European Spine Journal -