It is possible there are rare, but highly efficient, pollinators that were rarely observed during the sampling period, or were lumped together with a more frequently observed morphotype. An alternative explanation for the lack of an association between floral visitation and seed set is that higher plant diversity in urban and agricultural areas may decrease pollinator efficiency. Previous research has shown that invasive alien plants can have a negative effect on native plant communities by acting as attractors for pollinators, or decreasing pollinator efficiency by providing a wider range of resources for pollinators to visit, with the consequence that visitors transfer pollen from non con-specifics, potentially clogging stigmas and reducing pollination success. In this case, our target plant, yellow starthistle is indeed considered an invasive alien plant, but the hypothesis of it being in a novel diverse community could lead to a similar effect on the frequency and quality of pollination services that it receives. In sites where there are many other potential plants to visit and accompanying decreased floral fidelity leading to diverse pollen loads, one predicts decreased pollinator efficiency. Abundant sources of exotic plant pollen could occur in areas where there is a greater diversity of nearby plants for pollinators to visit. This explanation might account for the observation that shield-tipped small dark bees were negatively correlated with seed set. We selected yellow starthistle as the target plant for this study because of its ubiquitous distribution,planting gutter reliance on pollination, and its attraction for a wide set of visitors; it is also a highly invasive and undesirable plant.
Previous research on yellow starthistle has found that its invasion can be facilitated other non-native pollinator species such as the honey bee, Apis mellifera, and the starthistle bee, Megachile apicalis, which is included in the medium striped hairy belly bee morphotype. However, the abundance of bees in both of these 2 morphotypes were most closely associated with agricultural areas, which did not have the highest rates of seed set as would be predicted by visitation alone. Our results indicate clearly that bee visitation in human-altered landscapes can be higher than that in comparable natural areas, especially towards the end of the flowering season when there are few resources available in natural landscapes. Because the response of bee visitors to land use change depends on species specific requirements and these pollinators also have variable effects on plants, understanding the effect of land use change on pollination services requires knowledge not only of which pollinator groups shift to the human-altered landscapes, but also the rate of pollination that those groups have on the plant species in those landscapes. Future research will benefit from looking at a wider range of plants with a different range of target pollinators and that flower earlier in the year to better tease out these hypotheses. If the patterns of bee visitation and seed set that we observed are indeed consistent across other plant species, the novel plant communities created in these human-altered landscapes and the generalist bee species that are favored in such landscapes will lead to a reduction in overall pollination services.There is increasing evidence that alterations in the energy metabolism of cyst lining cells—especially increased glucose dependency and defects in fatty acid oxidation—may underlie the pathogenesis of autosomal dominant polycystic kidney disease.
Dietary interventions have been shown to be surprisingly effective in several polycystic kidney disease animal models, where they lead to a strong decrease in cyst growth. The positive effects of mild food reduction were most likely mediated by ketosis, as a ketogenic state—regardless of whether it was induced by a time-restricted diet, a ketogenic diet or a short term water fast —resulted in significantly inhibited cyst growth, fibrosis and PKD-associated signaling pathways in different animal models, even when the state of ketosis was only induced for a short period of time. Ketogenic dietary interventions are high-fat, low carbohydrate and moderate-protein diets which mimic a fasting state. KDIs have been used as an effective tool for the treatment of obesity and childhood epilepsy and could potentially be beneficial in several other diseases. A recent retrospective case series indicated safety, feasibility and positive effects of KDIs in patients with ADPKD for the first time. Most recently, the results of a 1- year behavioral weight loss study in obese ADPKD patients supported therapeutic feasibility of weight loss interventions and hinted towards possible positive effects such as slowing of kidney growth. However, no trials investigating the effects of ketosis per se in patients with ADPKD have been performed. Therefore, this proof-of-principle trial aimed to provide prospectively collected data on the short-term effects of KDIs in ADPKD patients.This trial was designed and conducted as a nonrandomized, non-blinded, single-center study at the University Hospital Cologne.Patients were recruited from the German ADPKD cohort or through the patients’ advocacy organization “Familiäre Zystennieren e.V.” If assessed eligible at the screening visit by medical history, physical examination and laboratory parameters, participants were enrolled after having obtained written informed consent.
The study was conducted in accordance with the Declaration of Helsinki and the good clinical practice guidelines by the International Conference on Harmonization.The study protocol will be provided upon request.Four study visits were conducted as part of the study . Each visit included blood and urine tests, bio-impedance measurements, measurements of anthropometric parameters, measurements of ketosis in finger stick blood, urine and breath, and an MRI abdomen for kidney and liver volumetry. Between V1 and V2 , patients continued to eat according to their usual dietary habits, i.e. a high-CHO diet, for a minimum of 13 and a maximum of 28 days. After V2, the KDI was started within 7 days and the KDI was finished by V3 . Patients could choose whether they wanted to achieve the ketogenic metabolic state by WF for 3 days or by a KD for 14 days. V3 was conducted after a maximum of 72 h upon termination of the diet. Between V3 and V4 , participants switched back to their standard ad libitum diet for a minimum of 20 and a maximum of 43 days. During I1 and I3, patients measured the extent of ketosis in breath, urine and fingertip blood at least three times, while in I2, during the KDIs, ketosis measurements were performed twice a day . Patients were provided with a diet diary for daily documentation of hunger,gutter berries well-being and potential additional foods consumed as well as results of the daily ketosis measurements. After completion of the KDIs, a dedicated questionnaire was used to assess feasibility and tolerability of the KDIs.Patients in the WF study arm limited oral intake to ad libitum amount of water and a low-salt broth once a day for a period of 3 days; in the KD study arm, patients consumed a very high-fat, low-CHO diet for a period of 14 days according to individual dietary plans. KD was based on a fat:protein:CHO ratio of 10:4:1 and calorie requirements were calculated individually for each patient with the MifflinSt. Jeor equation. Ten percent of the fat calories were provided as medium-chain triglycerides . Patients were supplied with the required food items. Patients in the KD group were advised to consume at least 20 kcal/kg body weight, but preferably 25 kcal/kg body weight daily. In addition, regular phone calls ascertained patients’ well-being and monitored adherence to the protocol. Patients in the KD arm were instructed to refrain from additional intake of non-ketogenic foods during the intervention. Both groups of patients received ketogenic snacks in case they desired food in between scheduled meals or experienced undesired symptoms during WF, respectively. Patients were advised to eat blueberries in case of blood beta-hydroxybutyrate levels >3.5 mmol/L or breath acetone levels > 40 p.p.m. and/or malaise or symptoms of ketosis .Baseline demographic and clinical data including comorbidities and medication were assessed at the screening visit. Anthropometric data were recorded at all study visits. Vital parameters were assessed at each study visit. Acetone concentrations were measured using a portable breath analyzer . BHB measurements in finger stick blood were performed with a portable ketone meter . Urine ketones were measured using urine dipsticks . The MRIs were performed by the in house Department of Radiology on a 1.5-T system .
For assessment of total kidney volume and total liver volume , kidney and liver boundaries of each patient were manually traced in axial T2 SPIR scans by a radiologist using Intellispace Discovery . The renal hilum was excluded from the kidney outline while the gallbladder and the main portal vein were excluded from the liver outline. A second reader was employed to additionally segment the kidneys and liver of each patient to estimate inter-reader variability in TKV and TLV. Both readers were blinded to patient information and previous tracing results. To evaluate cyst fraction in both organs, the T2 map was overlayed with each volume. Using Intellispace Discovery, each voxel within each volume was classified in either cystic or non-cystic depending on a cut-off value of 250 ms. This cut-off appeared to sufficiently differentiate tissue depending on its water content in a previous study and allows to approximate cyst fraction. The T1 mDIXON sequence which is included in our research protocol facilitates the assessment of fat content of liver tissue using chemical shift imaging. This allowed to check for potential fatty liver disease such as non-alcoholic fatty liver disease . Additionally, morphologic sequences were used to find potential fibrotic or cirrhotic changes of liver tissue. Clinical chemistry measurements on blood and urine samples were performed by the in-house central laboratory . Body composition was evaluated using a Tanita BC 418 MA scale .RESET-PKD was designed as a pilot trial analyzing pre-defined exploratory endpoints on feasibility and safety of short-term KDIs as well as their impact on TKV and TLV. Regarding TKV, we focused on the relative change in TKV between V2 and V3 . This analysis was complemented by the absolute and relative differences of TKV/height-adjusted TKV between study visits ; the same timepoints were compared regarding TLV, anthropometric parameters and blood pressure. To allow for the detection of potential safety signals a panel of blood and urine values including kidney function, lipids and liver values were examined , and feeling of hunger, discomforts and problems with general well-being were analyzed from the diet diary . The biochemical efficacy of the KDIs was assessed as follows: absolute and relative differences in self measurements of ketosis parameters . Most investigators agree that normal values for BHB on standard Western diets are 0.1–0.5 mmol/L. A non-linear relationship between acetone in breath and BHB in plasma has been described in adults. Therefore, we defined the cutoffs for the metabolic endpoints as follows: for the KD group: acetone level ≥10 p.p.m. or a BHB level ≥0.8 mmol/L in ≥75% of home measurements ; WF group: acetone level ≥10 p.p.m. or a BHB level ≥0.8 mmol/L in ≥75% of home measurements, or alternatively, a ketogenic state in at least one measurement in either method on 2/3 days during the WF. Feasibility was assessed using a questionnaire that contained 17 questions directly related to the KDI. Patients could rate each question on a scale from −4 to +4 with −4 representing low and +4 high feasibility . An average score of ≥0 was required to consider the KDI feasible. Both the metabolic endpoint and the feasibility endpoint had to be reached to meet the combined feasibility endpoint. The self reported feeling of hunger was recorded regularly in the study diary and linked to numbers from 1 to 4 . Stool, urine and blood samples were frozen for future analyses after patients had given specific informed consent for bio-banking.Both KDIs induced a significant weight loss . Loss of body water and loss of fat mass contributed equally . However, two patients in the KD group reported lower blood pressure values in their home measurements. In one of those patients, antihypertensive medication had to be paused upon the start of the ketogenicdiet due to orthostasis. All anthropometric parameters are provided in Supplementary data, Table S4.RESET-PKD is the first prospective interventional trial to combine exploratory analyses of metabolic efficacy , feasibility and efficacy of short-term controlled ketogenic metabolism in patients with ADPKD. Following the promising data on the beneficial effects of ketosis in PKD animal models, the present study was designed to include patients who could benefit most, i.e. rapid progressors.