deCLUTTER²⁺ – a pipeline to analyze calcium traces in a stem cell model for ventral midbrain patterned astrocytes

Astrocytes are the most populous cell type of the human central nervous system and are essential for physiological brain function. Increasing evidence suggests multiple roles for astrocytes in Parkinson’s disease, nudging a shift in the research focus, which historically pivoted around ventral midbrain dopaminergic neurons (vmDANs). Studying human astrocytes and other cell types in vivo remains challenging. However, in vitro-reprogrammed human stem cell-based models provide a promising alternative. Here, we describe a novel protocol for astrocyte differentiation from human stem cell-derived vmDAN-generating progenitors. This protocol simulates the regionalization, gliogenic switch, radial migration and final differentiation that occur in the developing human brain. We characterized the morphological, molecular and functional features of these ventral midbrain patterned astrocytes with a broad palette of techniques and identified novel candidate midbrain-astrocyte specific markers. In addition, we developed a new pipeline for calcium imaging data analysis called deCLUTTER2+ (deconvolution of Ca2+ fluorescent patterns) that can be used to discover spontaneous or cue-dependent patterns of Ca2+ transients. Altogether, our protocol enables the characterization of the functional properties of human ventral midbrain patterned astrocytes under physiological conditions and in disease.</p

Decision-making based on 3D printed models in laparoscopic liver resections with intraoperative ultrasound: A prospective observational study

OBJECTIVES: The aim of this study was to evaluate impact of 3D printed models on decision-making in context of laparoscopic liver resections (LLR) performed with intraoperative ultrasound (IOUS) guidance. METHODS: Nineteen patients with liver malignances (74% were colorectal cancer metastases) were prospectively qualified for LLR or radiofrequency ablation in a single center from April 2017 to December 2018. Models were 3DP in all cases based on CT and facilitated optical visualization of tumors\u27 relationships with portal and hepatic veins. Planned surgical extent and its changes were tracked after CT analysis and 3D model inspection, as well as intraoperatively using IOUS. RESULTS: Nineteen patients were included in the analysis. Information from either 3DP or IOUS led to changes in the planned surgical approach in 13/19 (68%) patients. In 5/19 (26%) patients, the 3DP model altered the plan of the surgery preoperatively. In 4/19 (21%) patients, 3DP independently changed the approach. In one patient, IOUS modified the plan post-3DP. In 8/19 (42%) patients, 3DP model did not change the approach, whereas IOUS did. In total, IOUS altered surgical plans in 9 (47%) cases. Most of those changes (6/9; 67%) were caused by detection of additional lesions not visible on CT and 3DP. CONCLUSIONS: 3DP can be helpful in planning complex and major LLRs and led to changes in surgical approach in 26.3% (5/19 patients) in our series. 3DP may serve as a useful adjunct to IOUS. KEY POINTS: • 3D printing can help in decision-making before major and complex resections in patients with liver cancer. • In 5/19 patients, 3D printed model altered surgical plan preoperatively. • Most surgical plan changes based on intraoperative ultrasonography were caused by detection of additional lesions not visible on CT and 3D model

LRP10 and α-synuclein transmission in Lewy body diseases

Autosomal dominant variants in LRP10 have been identified in patients with Lewy body diseases (LBDs), including Parkinson's disease (PD), Parkinson's disease-dementia (PDD), and dementia with Lewy bodies (DLB). Nevertheless, there is little mechanistic insight into the role of LRP10 in disease pathogenesis. In the brains of control individuals, LRP10 is typically expressed in non-neuronal cells like astrocytes and neurovasculature, but in idiopathic and genetic cases of PD, PDD, and DLB, it is also present in α-synuclein-positive neuronal Lewy bodies. These observations raise the questions of what leads to the accumulation of LRP10 in Lewy bodies and whether a possible interaction between LRP10 and α-synuclein plays a role in disease pathogenesis. Here, we demonstrate that wild-type LRP10 is secreted via extracellular vesicles (EVs) and can be internalised via clathrin-dependent endocytosis. Additionally, we show that LRP10 secretion is highly sensitive to autophagy inhibition, which induces the formation of atypical LRP10 vesicular structures in neurons in human-induced pluripotent stem cells (iPSC)-derived brain organoids. Furthermore, we show that LRP10 overexpression leads to a strong induction of monomeric α-synuclein secretion, together with time-dependent, stress-sensitive changes in intracellular α-synuclein levels. Interestingly, patient-derived astrocytes carrying the c.1424 + 5G &gt; A LRP10 variant secrete aberrant high-molecular-weight species of LRP10 in EV-free media fractions. Finally, we show that this truncated patient-derived LRP10 protein species (LRP10splice) binds to wild-type LRP10, reduces LRP10 wild-type levels, and antagonises the effect of LRP10 on α-synuclein levels and distribution. Together, this work provides initial evidence for a possible functional role of LRP10 in LBDs by modulating intra- and extracellular α-synuclein levels, and pathogenic mechanisms linked to the disease-associated c.1424 + 5G &gt; A LRP10 variant, pointing towards potentially important disease mechanisms in LBDs.</p

Metabolic and endocrine profiles and reproductive parameters in dairy cows under grazing conditions: effect of polymorphisms in somatotropic axis genes

<p>Abstract</p> <p>Background</p> <p>The present study hypothesized that GH-AluI and IGF-I-SnabI polymorphisms do change the metabolic/endocrine profiles in Holstein cows during the transition period, which in turn are associated with productive and reproductive parameters.</p> <p>Methods</p> <p>Holstein cows (Farm 1, primiparous cows, n = 110, and Farm 2, multiparous cows, n = 76) under grazing conditions were selected and GH and IGF-I genotypes were determined. Blood samples for metabolic/endocrine determinations were taken during the transition period and early lactation in both farms. Data was analyzed by farm using a repeated measures analyses including GH and IGF-I genotypes, days and interactions as fixed effects, sire and cow as random effects and calving date as covariate.</p> <p>Results and Discussion</p> <p>Frequencies of GH and IGF-I alleles were L:0.84, V:0.16 and A:0.60, B:0.40, respectively. The GH genotype was not associated with productive or reproductive variables, but interaction with days affected FCM yield in multiparous (farm 2) cows (LL yielded more than LV cows) in early lactation. The GH genotype affected NEFA and IGF-I concentrations in farm 1 (LV had higher NEFA and lower IGF-I than LL cows) suggesting a better energy status of LL cows.</p> <p>There was no effect of IGF-I genotype on productive variables, but a trend was found for FCM in farm 2 (AB cows yielded more than AA cows). IGF-I genotype affected calving first service interval in farm 1, and the interaction with days tended to affect FCM yield (AB cows had a shorter interval and yielded more FCM than BB cows). IGF-I genotype affected BHB, NEFA, and insulin concentrations in farm 1: primiparous BB cows had lower NEFA and BHB and higher insulin concentrations. In farm 2, there was no effect of IGF-I genotype, but there was an interaction with days on IGF-I concentration, suggesting a greater uncoupling somatropic axis in AB and BB than AA cows, being in accordance with greater FCM yield in AB cows.</p> <p>Conclusion</p> <p>The GH and IGF-I genotypes had no substantial effect on productive parameters, although IGF-I genotype affected calving-first service interval in primiparous cows. Besides, these genotypes may modify the endocrine/metabolic profiles of the transition dairy cow under grazing conditions.</p

The surface runoff of calcium, magnesium, iron, manganese, nitrogen and phosphorus from the upper Pasłęka river catchment

Celem badań było ustalenie wartości jednostkowych współczynników spływu makropierwiastków z terenów o różnych formach zagospodarowania w zlewni górnej Pasłęki i jej prawobrzeżnego dopływu Giłwy. Stwierdzono, że największe wartości współczynników spływu manganu oraz azotu i fosforu do wód uzyskano z obszarów użytkowanych rolniczo (110,8 kg Ca∙ha-1∙rok-1, 15,6 kg Mg∙ha-1 ∙rok-1, 0,5 kg Mn∙ha-1∙rok-1, 0,6 kg P∙ha-1∙rok-1, 14,3 kg N∙ha-1∙rok -1). Badania wykazały też wpływ terenów użytkowanych rolniczo, zarówno gruntów ornych, jak i użytków zielonych (łąki i pastwiska), na zwiększenie ładunku obszarowego wapnia i magnezu, w odniesieniu do których uzyskano największe współczynniki spływu z tych form użytkowania ziemi. Największą wartość współczynników spływu żelaza uzyskano dla obszarów leśnych (0,6 kg Fe∙ha-1 ∙rok-1).The aim of study was to determine values of unit coefficients of the surface runoff of macroelements from surfaces with different land management types, which was conducted in the catchment basin of the upper Pasłęka River and its right-bank tributary, the Giłwa River. The highest values of the coefficients describing the runoff of manganese as well as nitrogen and phosphorus to waters were obtained for a subcatchment comprising farmland (110.8 kg Ca∙ha-1·year-1, 15.6 kg Mg·ha-1·year-1, 0.5 kg Mn·ha-1·year-1, 0.6 kg P·ha-1·year-1, 14.3 kg N·ha-1·year-1 ). The study also pointed up the effect of agriculturally used areas, both arable land and grassland (meadows and pastures), consisting in increasing the non-point loads of calcium and magnesium, which scored the highest runoff coefficient values for the mentioned types of land use. The highest value of the iron runoff coefficient was calculated for forested areas (0.6 kg Fe·ha-1·year-1)

Feeling of satisfaction and being burdened with duties among informal care givers of the elderly according to the level of efficacy of care recipients

Wprowadzenie. Opiekunowie nieformalni, sprawujący opiekę nad starszym niesprawnym członkiem rodziny odczuwają nie tylko obciążenie, ale i satysfakcję z pełnionej funkcji. Cel pracy. Celem pracy jest określenie związku pomiędzy sprawnością podopiecznego a częstością odczuwanego przez opiekuna obciążenia obowiązkami oraz odczuwanej satysfakcji z pełnionej opieki. Materiał i metody. Badaniem objęto 64 opiekunów nieformalnych sprawujących nieodpłatną opiekę nad starszą osobą powyżej 65. roku życia, zamieszkujących gminę Zbuczyn Poduchowny oraz miasto Siedlce. W większości próbę badawczą wybrano stosując nieprobabilistyczną metodę tzw. kuli śnieżnej, dane od pozostałej części badanych zebrał lekarz psychiatra oraz 3 pielęgniarki środowiskowe. Materiał badawczy zgromadzono metodą sondażu diagnostycznego, w której wykorzystano autorski kwestionariusz ankiety oraz kwestionariusz oceny stanu podopiecznego wg skali Barthel (ADL). Wyniki. Opiekunowie, którzy odczuwali obciążenie zawsze lub często, zajmowali się osobami o niższym stopniu sprawności niż ci, którzy nigdy nie czuli obciążenia opieką (różnica wskaźnika sprawności podopiecznych z obu grup – 19 punktów w skali Barthel). Opiekunowie „zawsze” odczuwający satysfakcję – zajmowali się podopiecznymi o zbliżonym stopniu sprawności do tych, nad którymi opiekę sprawowały osoby, które nigdy nie czuły satysfakcji z pełnionej opieki, wypełnianej roli opiekuna (różnica wskaźnika sprawności podopiecznych z obu grup – 3 punkty w skali Barthel). Znaczną różnicę w poziomie sprawności podopiecznych zauważono między grupą opiekunów, którzy często czują satysfakcję oraz czasami (różnica wskaźnika sprawności podopiecznych z obu grup – 17 punktów w skali Barthel). Wnioski. Wzrasta deklarowana przez opiekuna częstość poczucia obciążenia opieką wraz ze spadkiem sprawności podopiecznego w zakresie ADL. Nie stwierdzono zależności pomiędzy sprawnością podopiecznego w zakresie ADL a poczuciem satysfakcji opiekuna ze sprawowanej opieki.Introduction. Informal caregivers providing care of an elderly, disabled family member feel not only the burden, but also satisfaction with the role performed. Objective. The objective of the study is determination of the relationship between the efficacy of care recipient and the frequency of the caregiver experiencing being burdened with duties and feeling of satisfaction with the care provided. Material and method. The study covered 64 informal caregivers providing free care of an elderly person aged over 65, living in the commune of Zbuczyn Poduchowny and the city of Siedlce. The study group was selected primarily by using the non-probabilistic method, the so-called ‘snowball’ method, data from the remaining part of the respondents were collected by a psychiatrist and 3 environmental nurses. Research material was collected by the method of a diagnostic survey, using a questionnaire constructed by the author, and the questionnaire of evaluation of care recipient status according to Barthel scale (ADL). Results. Caregivers who experienced burden always or frequently took care of persons with lower level of efficacy than those who never felt burdened with care (the difference in the indicator of efficacy of care recipients between both groups – 19 scores according to Barthel scale). Caregivers who ‘always’ felt satisfaction took care of persons with the degree of efficacy similar to that of caregivers who ‘never’ felt satisfaction with the care provided (the difference in the indicator of efficacy of care recipients between both groups – 3 scores according to Barthel scale). A considerable difference in the level of efficacy of care recipients was observed between caregivers who felt satisfaction ‘often’ and ‘sometimes’ (the difference in the indicator of efficacy of care recipients between both groups – 17 scores according to Barthel scale). Conclusions. The frequency of feeling being burdened with care by caregivers increases with the decrease in care recipient’s efficacy with respect to ADL. No relationship was found between care recipient’s efficacy regarding ADL, and caregiver’s feeling of satisfaction with the care provided