123903 research outputs found
Sort by
Impact of electrode surface texturing on hydrogen bubble dynamics during proton exchange membrane water electrolysis
No full textWater electrolysis systems are vital for sustaining human life during long-term space exploration missions. The main obstacle to the in-space operation of a water electrolysis system is the near-absence of buoyancy forces, impeding the detachment of hydrogen and oxygen bubbles from the electrodes and further complicating gas management, a crucial factor for efficient operation even in terrestrial applications. One of the most promising approaches to mitigate this problem is the micro- or nanostructuring of the electrode surfaces. Via surface structuring, the electrochemically active surface area can be enlarged and hydrophilicity increased, leading to easier detachment of gas bubbles. Additionally, bubble nucleation can be improved and bubble coalescence reduced. In this study, five pairs of laser-textured electrodes are manufactured, characterized and analyzed in terms of their performance and their influence on the bubble dynamics of the produced hydrogen gas. All textured electrodes achieve a performance enhancement over the unmodified surface (10% to 45% increase in current density for the same supply voltage). Gas production experiments prove that an increase in current density at a given voltage directly corresponds to a rise in production rate and, hence, in electrolysis performance. Significant differences in the bubble size distribution are observed on the different surfaces, as well as at different supply voltages. Distribution shapes and parameters (mean and standard deviation) remain mostly constant over time. Bubble rise velocities are significantly influenced by the entrainment of flow by the rising bubble plumes. Bubble growth after detachment is proven to be diffusion-controlled, and mainly determined by the degree of supersaturation close to the electrodes. This study proves that modification of the electrode surface morphology influences the performance of PEM systems by alteration of the bubble behavior during operation
Elucidating the significance of platelet proteasome function and its role in shaping the tumour microenvironment in vitro
No full textTrombociti imajo ključno vlogo pri regulaciji hemostaze, a so danes prepoznani tudi kot aktivni regulatorji imunosti, vnetja in napredovanja tumorjev. Kljub njihovim številnim biološkim vlogam pa ostajajo molekularni mehanizmi, ki uravnavajo aktivacijo trombocitov in njihovo sekretorno vedenje, pomanjkljivo opredeljeni. Nedavne raziskave so pokazale, da trombociti vsebujejo tako konstitutivni proteasom kot tudi imunoproteasom, ki ju običajno povezujemo predvsem z jedrnimi in imunskimi celicami. Ta doktorska disertacija preučuje, kako ti proteolitični sistemi oblikujejo biologijo trombocitov in vplivajo na trombocitno posredovane imunske in tumorske interakcije.
Klinično pomembnost delovanja proteasoma v trombocitih smo definirali s kvantifikacijo trombocitopenije pri bolnikih z diseminiranim plazmocitomom, zdravljenih z zaviralci proteasoma. Zaviranje proteasoma je pomembno povečalo pojavnost trombocitopenije, pri čemer je bilo tveganje odvisno od uporabljenega zaviralca in njegovega odmerka. V okviru mehanističnih raziskav smo zato okarakterizirali aktivnost proteasoma in imunoproteasoma v človeških trombocitih ter zaznali pomembne inter-individualne razlike v proteasomskih profilih, kar nakazuje, da sestava proteasoma prispeva k specifičnim razlikam med donorji v odzivnosti trombocitov. Za razliko od jedrnih celic trombociti ob izpostavitvi vnetnim citokinom ne povišajo proteinskega nivoja katalitičnih podenot imunoproteasoma, kar nakazuje, da je njihova proteasomska sestava določena že med megakariopoezo. Selektivno zaviranje katalitičnih podenot okrepi izločanje gostih granul, a ne vpliva na proces agregacije, kar dokazuje, da imunoproteasom natančno uravnava sekretorne odzive trombocitov, ne da bi pri tem ogrozil njihovo osnovno hemostatsko funkcijo. Proteomske analize so pokazale spremenjeno izločanje proteinov iz trombocitov, povezanih s presnovnimi in mitohondrijskimi potmi, kar kaže na vlogo imunoproteasoma pri uravnavanju trombocitne bioenergetike in medcelične komunikacije.
Selektivno zaviranje izbranih podenot proteasomov v trombocitih vodi v sproščanje dejavnikov, ki izrazito modulirajo vedenje celic raka dojk. Zlasti zaviranje podenote β1i sproži sekretorni profil, ki okrepi migracijo celic trojno negativnega raka dojk, poveča angiogeno signaliziranje ter zviša izražanje označevalcev značilnih za epitelijsko-mezenhimski prehod. Ti izsledki kažejo, da imunoproteasom trombocitov neposredno oblikuje tumorsko relevantne parakrine signale in lahko vpliva na metastatsko napredovanje.Platelets, traditionally recognized for their role in haemostasis, are now understood to actively regulate immune responses, inflammation, and tumour progression. Despite their broad functional repertoire, the molecular mechanisms governing platelet activation and secretion remain incompletely defined. Recent evidence has revealed that platelets contain both the constitutive proteasome and the immunoproteasome, proteolytic complexes previously thought to function primarily in nucleated immune cells. This dissertation investigates how these systems regulate platelet biology and influence platelet-driven immune and tumour interactions.
The clinical relevance of platelet proteasome function was assessed through a meta-analysis of thrombocytopenia in multiple myeloma patients treated with proteasome inhibitors. Proteasome inhibition significantly increased thrombocytopenia incidence, with risk dependent on inhibitor type and dosage, underscoring the hematologic consequences of targeting proteasomal pathways. Mechanistic studies in human platelets revealed substantial interindividual variability in proteasome and immunoproteasome activity, suggesting that proteasomal composition contributes to donor-specific differences in platelet responsiveness. Unlike nucleated cells, platelets do not upregulate immunoproteasome subunits in response to inflammatory stimuli, indicating that their proteasomal profile is established during megakaryopoiesis. Selective inhibition of immunoproteasome catalytic subunits modulated platelet secretion in a subunit-specific manner, enhancing dense-granule release without impairing aggregation. Proteomic analysis of activated platelet supernatants demonstrated altered secretion of proteins linked to metabolic and mitochondrial pathways, implicating the immunoproteasome in the regulation of platelet bioenergetics and intercellular communication.
Importantly, platelets with selectively inhibited immunoproteasome subunits released factors that markedly influenced breast cancer cell behaviour. Inhibition of the β1i subunit promoted migration of triple-negative breast cancer cells, increased angiogenic signalling, and induced markers associated with epithelial-mesenchymal transition. Collectively, this work establishes the platelet immunoproteasome as regulator of platelet secretion and signalling, with implications for immune regulation and tumour progression
Synthesis of 3(pyrrolidin-1-yl)pyridine-based heat shock protein 90 C-terminal domain inhibitors with anticancer activity
No full textProteini toplotnega šoka 90 (Hsp90) so šaperoni, ki z zagotavljanjem pravilnega zvijanja svojih proteinov klientov pomagajo pri proteostazi. Ker je ob tem njihovo izražanje močno povečano v rakavih celicah, predstavljajo obetavno tarčo v terapiji raka. Strukturno je Hsp90 dimer, katerega monomerna enota sestoji iz N-končne domene (NKD) z vezavnim mestom za ATP, srednje domene, kamor se vežejo proteini klienti, in C-končne domene (CKD), ki omogoča dimerizacijo Hsp90. Najprej so za načrtovanje izkoriščali predvsem N-končno ATP-vezavno mesto. Nekateri prvi zaviralci so v predkliničnih in kliničnih študijah izkazovali protirakavo delovanje, vendar so zaradi neželenih učinkov in sprožitve odziva toplotnega šoka njihov razvoj povečini opustili. Raziskave so se nato usmerile predvsem v zaviralce C-končne domene, za katere se je izkazalo, da ne izzovejo odziva toplotnega šoka.
V sklopu magistrske naloge smo na podlagi predhodno ovrednotene spojine TJD-337 sintetizirali nove zaviralce C-končne domene Hsp90 s pirolidinskim skeletom. Pripravili smo enajst končnih spojin, ki so vse vsebovale (S)-3,4-dikloro-N-(1-(6-formamidopiridin-3-il)pirolidin-3-il)benzamidni del, razlikovale pa so se v bazičnem centru in distančniku. Z uvedbo različnih bazičnih centrov smo proučili vpliv substituiranosti amina na jakost vezave. S spreminjanjem števila C-atomov v distančniku in ciklizacijo le-tega smo raziskali vpliv razdalje med bazičnim centrom in aromatom ter vpliv rigidnosti distančnika. Vse končne spojine smo z uporabo testa metabolne aktivnosti biološko ovrednotili na celicah raka dojke MCF-7. Najmočnejšo zaviralno aktivnost je imela spojina 21 (0,51 ± 0,07 μM) s ciklobutilnim distančnikom in primarnim aminom, ki ima izboljšano delovanje tudi v primerjavi s spojino TJD-337. S primerjavo rezultatov testiranja smo ugotovili, da sta za dobro zaviralno delovanje pomembna prisotnost bazičnega centra ter ustrezna razdalja med aromatsko skupino in bazičnim centrom na obeh koncih molekule. Dolg linearen distančnik (spojina 24) in odsotnost bazičnega centra (spojina 15) vodita v neaktivnost spojin, medtem ko je v primeru rigidiziranega distančnika (npr. spojina 21) aktivnost izboljšana. Dobljeni rezultati so spodbudni za nadaljnji razvoj pirolidinskih zaviralcev C-končne domene Hsp90 kot protirakavih spojin.Heat shock protein 90 (Hsp90) is a molecular chaperone that ensures the correct folding of its substrates and therefore plays an important role in proteostasis. Since its expression is strongly increased in cancer cells, Hsp90 represents a promising target in cancer therapy. The functional form of Hsp90 consists of two identical monomers, each with three domains: the N-terminal domain (NTD) with ATP-binding site, the middle domain, which binds client proteins and the C-terminal domain (CKD), responsible for dimerization of Hsp90. N-terminal domain inhibitors were discovered first. Although some of the first inhibitors showed anticancer activity in pre-clinical and clinical studies, they were later mostly abandoned due to their side effects and the induction of the heat shock response. The drawbacks associated with Hsp90 NTD inhibition led to the development of C-terminal domain inhibitors, which did not induce the heat shock response.
In the context of this master’s thesis, we synthesized novel Hsp90 C-terminal domain inhibitors, that were based on the previously evaluated compound TJD-337. All eleven newly prepared compounds contain (S)-3,4-dichloro-N-(1-(6-formamidopyridin-3-yl)pyrrolidin-3-yl)benzamide moiety and differ in the basic centre and spacer. By introducing varying basic centers, we examined the effect of amine substitution on the activity. By changing the number of C-atoms in the spacer and cyclizing it, we investigated the effect of the distance between the basic center and the aromatic system and the effect of the rigidity of the spacer. All of the final compounds were biologically assessed using the metabolic activity assay on the MCF-7 breast cancer cells. Compound 21 (0,51 ± 0,07 μM) with a cyclobutyl spacer and a primary amine, showed the most potent inhibitory activity. Comparing the results, we found out that the presence of a basic centre and the optimal distance between the benzamide group and the basic centre are important for inhibitory activity. The long, linear spacer in compound 24 and the absence of the basic center in compound 15 lead to inactivity of those compounds, while in the case of a rigidized cyclic spacer in compound 21, compounds activity is improved. The results are promising for the further development of Hsp90 C-terminal domain inhibitors and for the therapy of triple-negative breast cancer and other cancers
Continental contrasts in climate extremes that control tree fecundity
No full textIn 2023, more than half of olive harvests (Olea europaea) across Spain, Greece, and Türkiye were lost to drought. The same year late freeze destroyed 90% of the peach crop (Prunus persica) on the Georgia Piedmont and the apple crop (Malus domestica) in central New York, Vermont, and southern Quebec. Climate extremes now rank with the costliest threats to agriculture, but their role in forest recovery from diebacks that are happening globally is unknown for lack of tree fecundity estimates in forests. Tolerance of climate extremes could depend on past exposure but constrained by phylogenetic conservatism. We report a continental scale analysis of climate extremes and forest fecundity across North America and Europe showing that responses to late freeze and drought are happening now. Species differences are not explained by the traits typically included in ecological studies and they are weakly associated with phylogeny. Late freeze, that is, freezing temperatures that follow the onset of flower development in spring, is shown to be “normal” in North America, but not Europe, potentially explaining failed seed production due to delayed onset and the resultant shorter growing period by North American transplants dating back at least to the 18th century. Drought has thus far had the greatest impacts in dry forested regions, but here too, species differences are not explained by traditional trait values. If responses have been buffered from drought and late freeze by past exposure, acclimation and local adaptation prove inadequate as extremes intensify
Vital breath and the order of existence
No full textThis chapter explores the concept of breath (Qi 氣) in Chinese philosophy, emphasizing its intrinsic link to the relational order of the cosmos (Li 理). In this context, breath serves as a metaphor for the vital force Qi, the energy that animates all living beings. The concept of Qi transcends the physical act of breathing, representing an invisible yet pervasive energy that flows through all existence and connects individuals to the universe. The rhythmic pulsation of Qi, embodying both breath and vital energy, mirrors the natural cycles of existence— day and night, the seasons, and the interplay of life and death. However, Qi does not exist in isolation but is intimately tied to Li, the structural principle that governs and patterns the universe. In ancient Chinese philosophy, Qi initially referred to a standalone concept, denoting vapor, air, and the universal breath that sustains and permeates all forms of organic existence. Over time, it gradually became part of a binary conceptual pair as Chinese thought deepened its focus on the dynamic interaction between breath and body, substantiality and pattern, energy and structure. This shift affirmed that the vitality of breath (Qi) and cosmic order (Li) are mutually dependent and inseparable in the continuous unfolding of existence. 2 J. S. Rošker Just as human breath cannot manifest without the lungs to give it form and direction, Qi requires Li to shape its flow and coherence. By applying the so-called method of (post)comparative philosophical sublation, this chapter will illuminate the dialectical model of correlative complementarity, which underlies the integration of both concepts, revealing their tight interconnection and mutual interdependence
Control of a Robotic Cell Using a Programmable Logic Controller and the IO-Link Wireless Protocol
No full textDiplomsko delo obravnava vodenje robotske celice z uporabo sodobnih industrijskih komunikacijskih tehnologij. Novost v robotski celici je implementacija brezžične komunikacije IO-Link Wireless za krmiljenje robotskih prijemal, ki omogoča večjo fleksibilnost sistema ter zmanjšuje potrebo po obsežnem ožičenju. Celica vključuje merilni stroj, dva industrijska robota Yaskawa, dva transportna trakova ter industrijo kamero, pri čemer se vodenje celotnega sistema izvaja na programirljivem logičnem krmilniku.
Za nadzor, diagnostiko in prikaz stanja sistema je bil razvit uporabniški vmesnik, ki omogoča enostavno spremljanje in upravljanje sistema v realnem času.
Celica je bila uspešno postavljena in preizkušena v praksi, kar potrjuje zanesljivost brezžične komunikacije ter učinkovitost integriranega pristopa. Naloga ponuja celovit vpogled v načrtovanje sistema za vodenje in prispeva k širši uporabi tehnologije IO-Link Wireless v industrijskem okolju.The bachelor’s thesis addresses the control of a robotic cell using modern industrial communication technologies. The novelty in the robotic cell is the implementation of IO-Link Wireless communication for controlling robotic grippers, which enables greater system flexibility and reduces the need for extensive cabling. The cell includes a measuring machine, two Yaskawa industrial robots, two conveyor belts, and an industrial camera, while the control of the entire system is carried out on a programmable logic controller.
For monitoring, diagnostics, and visualization of the system status, a user interface was developed that enables simple monitoring and management of the system in real time.
The cell was successfully set up and tested in practice, which confirms the reliability of wireless communication and the efficiency of the integrated approach. The thesis offers a comprehensive insight into the design of a control system and contributes to the wider use of IO-Link Wireless technology in an industrial environment
Fast evaluation of central moments for non-Gaussian random loads in vibration fatigue
No full textIn vibration fatigue analysis, spectral methods are used to evaluate the fatigue damage of structures experiencing random vibrations. Spectral methods fail under non-Gaussian and non-stationary loading conditions and various solutions have been proposed. Correction coefficients are promising and depend on the kurtosis and skewness of the system’s response, which requires extensive time-domain analyses. Performing time-domain analysis undermines the computational efficiency of spectral methods. The present manuscript proposes a modal decomposition-based approach to numerically efficiently compute the central moments required to obtain the kurtosis and skewness. The proposed method is numerically validated on a structure subjected to non-Gaussian random loads. The proposed method demonstrates results identical to the standard approach, showing a reduction in computation time of around two orders of magnitude. This extends the applicability of spectral methods in conjunction with correction coefficients for numerical estimation of fatigue damage in the frequency domain even in the case of non-Gaussian loadings
Progress in overtourism discourse measured through the well-being-centred sustainable tourism paradigm
No full textFunctional evaluation of FUBP3, MPP7 and ANAPC1 genes in the development of osteoporosis and osteoartrosis
No full textOsteoporoza (OP) je pogosta sistemska bolezen skeleta, za katero sta značilni zmanjšana mineralna kostna gostota (BMD) in povečano tveganje za zlome. Nasprotno je osteoartroza (OA) primarno bolezen sklepov, povezana s povišano BMD v subhondralni kosti, zato v raziskavah predstavlja obratno patologijo oziroma nasprotni fenotip OP. OP sodi med kompleksne genetske bolezni, pri katerih na razvoj vplivajo številni okoljski in genetski dejavniki z majhnimi učinki, kar otežuje njihovo identifikacijo in vrednotenje. Pomemben napredek so prinesle vsegenomske asociacijske študije (GWAS), ki so identificirale številne lokuse in gene, povezane z BMD in zlomi, vendar brez funkcionalne potrditve njihove vloge.
V svoji disertaciji smo se osredotočili na ocenjevanje funkcije treh genov – FUBP3, ANAPC1 in MPP7, saj so lokusi s temi geni pri GWAS izkazali močno značilno (p<10-8) povezanost z BMD in zlomi (FUBP3: p = 3,4 × 10?22, ANAPC1: p = 1,5 × 10?? in MPP7: p = 2,4 × 10?16). Zasnovali smo večnivojski eksperimentalni pristop, ki je vključeval: analizo izražanja genov v kostnem in mišičnem tkivu bolnikov z OP, OA ter kontrolnih oseb, spremljanje izražanja med diferenciacijo mezenhimskih stromalnih/matičnih celic (MSC) v osteoblaste, adipocite in miocite ter eksperimentalno manipulacijo izražanja (utišanje oziroma izbitje gena) v celičnih modelih.
V kostnem tkivu bolnikov z OP smo dokazali znižano izražanje vseh treh genov, pri OA pa le gena FUBP3. V mišičnem tkivu je bilo izražanje pri OP prav tako znižano, kar nakazuje pleiotropno delovanje teh genov. Med osteogeno diferenciacijo MSC se izražanje FUBP3 in ANAPC1 značilno zviša, kar potrjuje njuno vlogo v osteogenezi. Pokazali smo, da utišanje ANAPC1 vpliva na zgodnje faze osteogeneze, izbitje MPP7 povzroči popolno blokado mineralizacije, izbitje FUBP3 pa vodi v oslabljeno mineralizacijo.
Naši rezultati prvi potrjujejo vlogo genov FUBP3, ANAPC1 in MPP7 v kostni biologiji pri človeku ter njihovo povezavo z OP in OA. Raziskava prispeva k razumevanju genetske osnove OP in predstavlja korak k prepoznavanju novih biomarkerjev ter razvoju ciljnih terapij za personalizirano zdravljenje osteoporoze.Osteoporosis (OP) is a common systemic skeletal disorder characterized by reduced bone mineral density (BMD) and an increased risk of fractures. In contrast, osteoarthrosis (OA) is primarily a joint disease associated with elevated BMD in the subchondral bone and therefore represents an opposite pathology, or a reverse phenotype, of OP in research studies. OP is a complex genetic disease influenced by numerous environmental and genetic factors with small individual effects, which makes their identification and evaluation challenging. Significant progress has been achieved through genome-wide association studies (GWAS), which have identified multiple loci and genes associated with BMD and fractureshowever, these findings lack functional validation of gene roles.
In this dissertation, we focused on evaluating the function of three genes – FUBP3, ANAPC1, and MPP7 – as GWAS loci containing these genes showed strong genome-wide significant associations (p < 10⁻⁸) with BMD and fractures (FUBP3: p = 3.4 × 10⁻²²ANAPC1: p = 1.5 × 10⁻⁹MPP7: p = 2.4 × 10⁻¹⁶). We designed a multi-level experimental approach that included: analysis of gene expression in bone and muscle tissue of patients with OP and OA as well as control subjects, monitoring of gene expression during differentiation of mesenchymal stromal/stem cells (MSC) into osteoblasts, adipocytes, and myocytes, and experimental manipulation of gene expression (silencing or knockout) in relevant cellular models.
In bone tissue of patients with OP, the expression of all three genes was decreased, whereas in OA only FUBP3 expression was reduced. In muscle tissue, gene expression was also decreased in OP, indicating pleiotropic functions of these genes. During osteogenic differentiation of MSC, expression of FUBP3 and ANAPC1 significantly increased, confirming their involvement in osteogenesis. We demonstrated that silencing of ANAPC1 affects the early stages of osteogenesis, knockout of MPP7 results in complete inhibition of mineralization, and knockout of FUBP3 leads to impaired mineralization.
Our results provide the first evidence of the roles of FUBP3, ANAPC1, and MPP7 in human bone biology and their association with OP and OA. This study contributes to the understanding of the genetic basis of osteoporosis and represents an important step toward the identification of novel biomarkers and the development of targeted therapies for personalized treatment of osteoporosis
Study of drug supersaturation and precipitation mechanisms in vitro under simulated in vivo conditions
No full textOne of the key processes for drug absorption after oral administration is drug dissolution in the gastrointestinal (GI) tract fluids. Due to the use of complex drug delivery systems or changes in the physiological properties of the GI tract, supersaturation and precipitation of drugs can also occur. In vitro models that simulated various physiological parameters were developed within the doctoral dissertation to investigate drug dissolution, supersaturation, and precipitation. Using a pH shift method, drug precipitation was studied in a simple precipitation model with an innovative approach to simultaneously monitor the particle size distribution and concentration of the dissolved drug. Furthermore, we also developed several biorelevant models with separately simulated selected GI tract parameters: in vivo pH profiles along the GI tract, the gastric emptying kinetics of co-ingested water, and a combination of pH value and lipid concentration representing the gastric conditions at different times after ingestion of a meal. The sample preparation procedure and HPLC method were also developed in experiments with simulated fed state conditions to determine the amount of dissolved drug in the whole sample and its partitioning between the lipid and aqueous phases of the medium. Typical changes in the particle size distribution were observed in the precipitation model. The particles also increased the measured absorbances in a wavelength-dependent or wavelength-independent way concerning the particle size, thus also affecting the calculated amount of the dissolved drug. A distinctive effect of the simulated physiological parameters in biorelevant models on drug behaviour was observed, however, the solubility of drugs in tested media controlled the influence of other simulated parameters on drug behaviour. Since supersaturation and precipitation also occurred in biorelevant models, the identified precipitation mechanisms from the precipitation model were successfully translated into biorelevant models. This enabled a better understanding of the complex processes of model drug behaviour after oral administration, including drug dissolution, supersaturation, and precipitation.One of the key processes for drug absorption after oral administration is drug dissolution in the gastrointestinal (GI) tract fluids. Due to the use of complex drug delivery systems or changes in the physiological properties of the GI tract, supersaturation and precipitation of drugs can also occur. In vitro models that simulated various physiological parameters were developed within the doctoral dissertation to investigate drug dissolution, supersaturation, and precipitation. Using a pH shift method, drug precipitation was studied in a simple precipitation model with an innovative approach to simultaneously monitor the particle size distribution and concentration of the dissolved drug. Furthermore, we also developed several biorelevant models with separately simulated selected GI tract parameters: in vivo pH profiles along the GI tract, the gastric emptying kinetics of co-ingested water, and a combination of pH value and lipid concentration representing the gastric conditions at different times after ingestion of a meal. The sample preparation procedure and HPLC method were also developed in experiments with simulated fed state conditions to determine the amount of dissolved drug in the whole sample and its partitioning between the lipid and aqueous phases of the medium. Typical changes in the particle size distribution were observed in the precipitation model. The particles also increased the measured absorbances in a wavelength-dependent or wavelength-independent way concerning the particle size, thus also affecting the calculated amount of the dissolved drug. A distinctive effect of the simulated physiological parameters in biorelevant models on drug behaviour was observed, however, the solubility of drugs in tested media controlled the influence of other simulated parameters on drug behaviour. Since supersaturation and precipitation also occurred in biorelevant models, the identified precipitation mechanisms from the precipitation model were successfully translated into biorelevant models. This enabled a better understanding of the complex processes of model drug behaviour after oral administration, including drug dissolution, supersaturation, and precipitation