Saccharomyces cerevisiae, highlighting the caffeine synergy with external cell stressors, such as irradiation or exposure to various chemical hazards, including cigarette smoke or chemical carcinogens. The review also focuses on the importance of caffeine-related yeast phenotypes used to resolve molecular mechanisms involved in cell signaling through conserved pathways, such as target of rapamycin (TOR) signaling, Pkc1-Mpk1 mitogen activated protein kinase (MAPK) cascade, or Ras/cAMP protein kinase A (PKA) pathway.Caffeine–a methylxanthine analogue of the purine bases adenine and guanine–is by far the most consumed neuro-stimulant, being the active principle of widely consumed beverages such as coffee, tea, hot chocolate, and cola. While the best-known action of caffeine is to prevent sleepiness by blocking the adenosine receptors, caffeine exerts a pleiotropic effect on cells, which lead to the activation or inhibition of various cell integrity pathways. The aim of this review is to present the main studies set to investigate the effects of caffeine on cells using the model eukaryotic microorganism  Caffeine  is the best-known chemical constituent of coffee and one of the most widely consumed and socially accepted natural stimulants. As an important constituent of coffee, but also of other largely-consumed beverages such as tea, chocolate, and cola-like drinks, caffeine is by far the most ingested methylxanthine, along with the less representative theophylline  and theobromine . Caffeine is also widely used as an important ingredient of various medicine and non-prescription drugs , sports and energy drinks, nutritional supplements, and cosmetics. The scientific literature dealing with the biological effects of caffeine is vast, revealing a large amount of evidence on both the beneficial and deleterious effects, on indications and contraindications, on adverse effects and toxicity, etc. The action of caffeine at the cellular level has been intensively investigated, and there are three fundamental mechanisms which are universally recognized: intracellular mobilization of calcium, inhibition of phosphodiesterases, and antagonism at the level of adenosine receptors [Caffeine belongs to the purine alkaloid family closely linked with the bases adenine and guanine , which aSaccharomyces cerevisiae as a model for the eukaryotic cell. A model organism is used in scientific research for various reasons: simplification of the biological context, overcoming ethical and experimental constraints, elimination of redundancies, the establishment of a framework for development and optimization of analytical methods, etc. Importantly, a model organism has to be representative of a larger class of living beings [S. cerevisiae, a relatively simple unicellular eukaryote, has emerged as a versatile and robust model organism to study the fundamental factors that determine eukaryotic cell biology [S. cerevisiae is most utilized by the research community due to its amenability to genetic studies, comprehensive genome annotation [S. cerevisiae is an invaluable tool in genomic studies, resistance profiling, metabolome studies, and metabolic engineering [S. cerevisiae offers insights into the complex mechanisms underlying the sensing and response to the external conditions, including exposure to a plethora of synthetic and natural chemical compounds, such as caffeine. S. cerevisiae is generally responsive to caffeine, as it was uncovered that this substance affects yeast cell growth and morphology, DNA repair mechanisms, intracellular calcium homeostasis, and cell cycle progression [In this review, we present some of the studies set to unravel the caffeine mechanisms of action using g beings . S. cere biology . S. cerenotation , and a hnotation . Additioineering ,10,11,12gression .S. cerevisiae to unravel some potential effects of caffeine on the eukaryotic cells, with a focus on caffeine transport in yeast cells, caffeine influence on cells exposed to irradiation, caffeine interaction with target of rapamycin (TOR) and cell wall integrity pathways, and caffeine influence on the lifespan of the cells. This paper provides an overview on the studies that used S. cerevisiae cells has not been investigated in detail. Being non-essential, it is expected that caffeine would be carried into the cell by a non-specific transporter, such as purine permease. In S. cerevisiae, FCY2 encodes for a purine-cytosine permease, which mediates purine  and cytosine accumulation [fcy2Δ knock-out strain cannot accumulate caffeine from the medium [FCY21 and FCY22, which belong to the same family as FCY2. Nevertheless, despite of the nucleotide sequence similarity, neither FCY21 nor FCY22 can complement FCY2 absence [The effects of caffeine on cells are pleiotropic, causing delays to cell cycle progression; changes in cell morphology; and in high doses, cytotoxicity. Due to structural similarity to nucleotides , it has mulation . Fcy2 mae medium . In the  absence , and theSNQ2 was identified as a caffeine-resistance gene by screening a genomic library of S. cerevisiae in a multicopy vector. Multicopy of PDR5 also conferred resistance to caffeine but to a lower extent compared to SNQ2 [S. cerevisiae cells [S. cerevisiae strain, it was found that caffeine resistance could be gained generally by overexpression of pleiotropic drug resistance genes. The study identified a mutation in PDR5 but also in PRD1, which encodes the transcription factor which regulates PDR5 and SNQ2 expression, indicating that resistance to caffeine can be correlated with an efficient and active system of extrusion from the cell [BFR1 from Schizosaccharomyces pombe was expressed into S. cerevisiae, resulting in enhanced caffeine resistance, suggesting that ABC-transporters can be an efficient way to reduce caffeine toxicity in heterologous systems [Caffeine efflux into the extracellular space is better understood, and it is ensured by two ATP-binding cassette (ABC)-transporters responsible for the multidrug resistance in yeast, i.e., Snq2 and Pdr5. In fact, Snq2 was firstly described as the transporter responsible for caffeine detoxification, when  to SNQ2 . Pdr5 isae cells . Using ethe cell . An ABC- systems . In mamm systems  and whil systems ,22,23,24S. cerevisiae by overexpression of HSE1 , RTS3 , and SD budding ). None o budding .S. cerevisiae can be used to develop a model to study the toxic effects of various substances [S. cerevisiae cells when pure caffeine was used [rad1 and rad6 mutants of S. cerevisiae, whereas no effect was observed in the rad52 mutant [S. cerevisiae has been also used as a model system to explore the toxicity of the antitumoral agent 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU), when caffeine showed some effect in enhancing BCNU toxicity by decreasing both the mutagenic and the recombinogenic potential of the drug [The pleiotropic effect of caffeine on bstances . It was bstances . In linewas used . Caffein2 mutant . The intthe drug . A similthe drug .An exogenous substance with biological activity is expected to exert some effect at the plasma membrane level, and the ability of caffeine to block calcium entry into the yeast cell is one of such effects . CaffeinS. cerevisiae cells is a suitable organism to study the effect of radiation upon the eukaryotic cell and also a convenient platform to identify chemicals that alter this interaction. Some early studies suggested that caffeine potentiates the biological effects of radiation and chemical mutagens in a variety of organisms, including S. cerevisiae [revisiae .S. cerevisiae cells treated with caffeine show a significant increase in radio-sensitivity to various UV doses [UV doses , and it UV doses . These eUV doses . Later, UV doses ,43.+cdc colonies under UV irradiation. In S. cerevisiae, the CDC8 gene encodes for a thymidylate kinase involved in DNA replication, also required when UV irradiation induces gene conversion and gene mutation events and induction of +cdc colonies. Inhibition of DNA replication by caffeine diminishes the formation of +cdc colonies, indicating that the latter arises as a result of errors in DNA replication [Caffeine was also shown to inhibit some DNA repair mechanisms, reducing the generation of lication .S. cerevisiae cells was also studied for γ-irradiation. A slight caffeine-sensitizing effect was found for the rad5l and rad54 mutants, which show defects in the repair of X-ray induced damage [The effect of caffeine on irradiated d damage . The resd damage .S. cerevisiae cells was also analyzed by a single-cell gel electrophoresis of spheroplasts. By monitoring the γ-radiation-induced DNA damage, repair, and radioprotection, this study indicated a radioprotective effect of caffeine in a dose-dependent manner [The DNA damage induced by γ-irradiation of t manner .S. cerevisiae cells have two different physiological states, aerobic and anaerobic. Cells anaerobically grown do not have functionally active mitochondria, and the energy is generated only through glycolysis. It was shown that caffeine acted like a radioprotector against γ-radiation only in the case of yeast cells grown aerobically, in the presence of oxygen. The radioprotection offered to aerobically-grown cells did not influence the recovery process through biosynthetic reparatory ways, as caffeine did not influence the DNA repair process directly. Rather, the caffeine radioprotective phenotype observed involved scavenging of the reactive oxygen species produced by irradiation [The adiation . In contadiation .60Co-generated γ-radiation or of 4-nitroquinoline 1-oxide (4-NQO) exposure. The results indicated that caffeine decreased γ-radiation-induced gene conversion frequencies. In contrast, caffeine was found to increase the induced gene conversion frequency in cells treated with 4-NQO, suggesting that the repair processes following γ-irradiation or 4-NQO treatment involve different pathways [The effect of caffeine was also monitored in relation with the mutagenic action of pathways .S. cerevisiae, the response to DSB is controlled by DNA damage checkpoint signal-transduction pathways, which include the redundant protein kinases Mec1 and Tel1, members of the family of phosphatidyl inositol 3 (PI3) kinases [Irradiation is often linked to DNA damage events; therefore, it was natural to investigate the effect of caffeine on cell response to DNA damage. In yeast cells, caffeine was shown to inhibit some checkpoint kinases involved in DNA double-strand breaks (DSB) repair. DSB are highly deleterious events that may lead to chromosomal abnormalities, cell death, and cancer, and repair of chromosome breaks occurs by several highly conserved pathways . In S. c kinases ,51,52, wInteraction between yeast cells and caffeine was used to demonstrate that Mec1/Tel1-dependent intra-S-phase checkpoint activation inhibits Rad52 foci formation, which occurs as a response to replication forks collapsing . InductiUpon activation, Mec1 and Tel1 also act directly on chromatin by phosphorylating histone H2A on seryl-129 residue to yield H2A-S129 . When caS. cerevisiae, even the presence of a single DSB produces G2/M arrest, before anaphase [srs2Δ cells, supporting the idea that caffeine has the ability to override DNA damage checkpoints. Even though the cells lacking Srs2p helicase apparently completed DNA repair after caffeine treatment, the cells failed to recover, proving that Srs2p is required to turn off the DNA damage checkpoint. It was observed that inactivation of the checkpoint restores the viability of most srs2Δ cells, indicating that the cause of lethality of these mutant cells is the incapacity to turn off the checkpoint after the completion of DNA repair [In eukaryotes, DNA damage triggers the DNA damage checkpoint, causing cells to become blocked in cell cycle progression . In S. canaphase . Sometimanaphase . Srs2 isA repair .S. cerevisiae, KIN3 was identified as a gene that encodes for a structural homolog of NIMA serine-threonine kinase required in Aspergillus nidulans for DNA damage response and in the regulation of G2/M phase progression [S. cerevisiae cells that were either deleted for KIN3 or were overexpressing it had no detectable growth phenotypes, but it was noticed that caffeine abolished KIN3 expression induced by genotoxic agents, such as methyl methanesulfonate (MMS), cisplatin and doxorubicin, indicating that Kin3-activating signal is mediated by the caffeine-sensitive pathways. As caffeine can inhibit the DNA damage checkpoint transducers Mec1 and Tel1 [Another kinase targeted by caffeine in yeast is Kin3 kinase. In gression ,60. S. cand Tel1 ,61, it wand Tel1 .TOP3, whose deletion in S. cerevisiae causes hyperrecombination, meiotic defects, sensitivity to genotoxic agents, and poor growth due to accumulation of S/G2 DNA [TOP3 after exposure to mutagen MMS, it was observed that caffeine-treated cells successfully traverse S phase, while caffeine non-treated cells failed to show any significant recovery and remained with a mid-S DNA content, suggesting that a persistent checkpoint-mediated cell cycle delay leads to the impaired S-phase progression that can be overridden by the addition of caffeine [Topoisomerases are highly conserved proteins, required for many aspects of DNA metabolism. In yeast, DNA Topoisomerase III is encoded by gene S/G2 DNA . In a cocaffeine .S. cerevisiae cells was also used for studies on Ribonucleases H [RNH1 and RNH2 genes, respectively [RNH1, RNH2, or both, and it was noticed that the absence of RNase H1 in a strain that has an active RNase H2 diminishes the deleterious effects of caffeine and that in caffeine-treated cells, the un-degraded RNA-DNA hybrids influence DNA synthesis by damaging or perturbing the cell cycle [Interaction of caffeine with leases H . Ribonucectively . The effll cycle .Evolutionarily conserved target of rapamycin (TOR) kinase is a major regulator of cell growth and metabolism in response to a broad set of environmental signals and stress conditions. Because its defects were noted to be involved in disorders such as cancer, neurological, metabolic, inflammatory, and autoimmune diseases, as well as in ageing ,68,69, TS. cerevisiae has been very useful as model organism for understanding the role of TOR signaling in the regulation of cell growth and aging [The TOR pathways regulate the cellular growth under normal conditions, by stimulating ribosome biogenesis and by controlling the precursors for amino acids and other nitrogenous molecules’ synthesis a. Under nd aging ,76, and nd aging . Rapamycnd aging .The macrolide drug rapamycin is a macrocyclic lactone used as immunosuppressive and anti-proliferative antibiotic which inhibits TORC1 . In the In yeast, TORC1 contains kinases Tor1 or Tor2, as well as several additional proteins, including Kog1, Lst8, and Tco89; TORC2 contains Tor2 as well as Lst8, Avo1-Avo3, and Bit61  Figure b. In mamReinke et al. 2006  were amoS. cerevisiae uses a broad spectrum of compounds as nitrogen sources, and NCR is a physiological response when cells are grown under normal conditions, and preferred nitrogen sources are used  [The yeast cells treated with caffeine or rapamycin have a transcriptional profiling that proves the inhibition effect of TOR signaling on a broad array of genes associated with a wide range of cellular growth-related functions and also with stress and autophagy-related genes ,90. Notautamine) . In cellutamine) . In thisutamine) .S. cerevisiae, the Rho family has six members, Rho1 to Rho5 and Cdc42. Mutants with RHO5 gene deleted (rho5Δ) had a higher resistance to caffeine, in contrast to the Slt2 mitogen activated pathway kinase (MAPK) mutants, which were highly sensitive to caffeine, indicating a role for RHO5 in the down regulation of the Slt2-MAPK pathway. This special behavior was explained by the fact that Rho5 acts as an off-switch for the MAPK cascade, which differentiates between MAPK-dependent and independent functions of Pkc1, a prototypic member of the protein kinase C superfamily and the main effector of Rho1 [Rho-family GTPases are key regulators involved in many eukaryotic cell functions  and represent a core component of the TORC1 pathway ,100,101. of Rho1 . Rho1 is of Rho1 . In stre of Rho1 ,104. Upo of Rho1 . It was Δgtr1 and Δgtr2 have the similar caffeine-sensitive phenotype [N-terminus is required for Gtr1p–Gtr2p heterodimer formation but not for complex formation with Ego1p, a vacuolar membrane protein. Upon caffeine treatment, the amount of free Gtr1p increases, while it decreases in the protein complexes. Likewise, free Gtr2p is increased by caffeine but the amount form bound in the high molecular weight complexes remains unaffected, indicating that Gtr1p and Gtr2p are necessary for caffeine resistance and that caffeine treatment released Gtr1p from the Gtr1p–Gtr2p complex [The Ras-family-GTPase and its homologs  mediate henotype , in conchenotype ,91. Morehenotype , the EGOhenotype ,112. The complex .In a study over the sensitivity of TORC1 during the rapamycin treatment, it was revealed that the cells recovered efficiently from treatment with saturating concentration of rapamycin alone, as well as with the caffeine alone, and that caffeine is a selective inhibitor of rapamycin-insensitive proliferation; at the same time, the rapamycin-caffeine co-treatment followed by recovery in the presence of caffeine, induced a strong recovery defect . These oS. cerevisiae. Study of Ylr257w would be particularly informative since it was the only protein whose removal from the constructed network blocked the signal transduction to the TORC1 effector kinase Npr1 [Some new components of TOR signaling were recently identified following direct and specific inhibition of TOR signaling by caffeine and rapamycin using a network-based multi-omics integrative analysis that employed data from transcriptomics, interactomics, and regulomics sources in yeast ,112,113.ase Npr1 ,111.ptc1 mutants. Ptc1 is a 2C phosphatase isoform, member of the 2C phosphatase family; a connection with TOR pathways is not shared by most members of the family. Ptc1 is required for normal Gln3 and Msn2-mediated transcriptional responses and nuclear localization [ptc1 mutants exposed to rapamycin and caffeine, the translocation of Gln3 and Msn2 to the nucleus is prevented and also the dephosphorylation of the Npr1 kinase. At the same time, the overexpression of other isoforms (such as PTC2 or PTC3) did not confer tolerance to rapamycin, and ptc1 ptc6 double mutant were more sensitive to both rapamycin and caffeine, suggesting the role of both phosphatases in the signaling of TOR pathway [A functional link between Ptc1 and the TOR pathway was established due to the rapamycin and caffeine sensitivity of yeast lization . In yeas pathway .At the level of the general amino acid control (GAAC) and TOR pathways, the cellular stress response is regulated by the amino acylation status of the cellular tRNA pool, which directs the transcriptional regulation of gene expression in response to nutritional stresses . Under nIn yeast, twelve lysine methyltransferases that modify translational elongation factors and ribosomal proteins were identified. Among them, five  are specific to elongation factor 1A (EF1A), the protein responsible for bringing aminoacyl-tRNAs to the ribosome. It was demonstrated that loss of EF1A methylation is not essential to cell viability but leads to a decrease in growth rates under caffeine and rapamycin treatment. These findings suggested that EF1A interacts with the TORC1 pathway and that Efm methyltransferases are devoted to the modification of EF1A, finding no evidence for the methylation of other substrates in the yeast cell .hhy mutants), all the hhy mutants revealed severe sensitivities to caffeine and rapamycin, suggesting an interaction between the identified genes in TOR kinase pathway [The mammalian lysosome has an analogue in yeast, the vacuole, a membrane-bounded organelle. The yeast vacuole contains an acidic environment due to vacuolar hydrolases that degrade structural debris macromolecules and waste products . In a ge pathway ,119.S. cerevisiae confers cell shape and protection against harsh environments [Candida spp., Cryptococcus neoformans, Aspergillus spp., Pneumocystis carinii, or Histoplasma capsulatum [S. cerevisiae, it was observed that sensitivity to caffeine can be correlated with defects in the CWI pathway and that caffeine activates CWI signaling, when the stability of the cell wall can be monitored in terms of response to osmotic or thermal stress [C-terminus at Ser423 and Ser428 residues independently of the standard dual phosphorylation associated with MAPK activation; nevertheless, these phosphorylations are dependent on the DNA damage checkpoint kinases, Mec1/Tel1 and Rad53 [PRS (phospho ribosylpyrophosphate synthetase) genes, in particular those lacking the Prs1/Prs3 minimal functional unit [PRS1, there is a correlation between caffeine sensitivity and increased basal expression of Rlm1, the transcription factor which is an important component of the PKC-mediated MAPK pathway involved in the maintenance of CWI [Investigations regarding the interaction between caffeine and yeast cells demonstrated the existence of additional caffeine targets, including components of cell wall integrity (CWI) pathways . The celronments . It is fpsulatum . The CWIpsulatum . Rho1p Gpsulatum , and actpsulatum . A plethpsulatum . In S. cl stress . It was nd Rad53 . Other snal unit . In altee of CWI .S. cerevisiae genome. Among these, YNL313c, renamed EMW1 , proved to be essential for the maintenance of CWI, and the mutants lacking EMW1 showed sensitivity to diverse stressor compounds, including caffeine [rim21Δ (ynl294c) showed a moderate hypersensitivity to caffeine owed to a low compensatory response of the cell wall, indicated by the almost complete absence of Slt2 phosphorylation and the modest increase in chitin synthesis after calcofluor treatment [The loss of function of other proteins involved in CWI can also be related to caffeine sensitive phenotypes. For example, there are six proteins that have the tetratricopeptide repeat (TPR) domain (mediates protein–protein interaction), which are encoded by six essential genes in the caffeine . Moreovereatment .S. cerevisiae genes (approximately 6000 genes), the yeast has 117 protein kinase (PKase) and 32 protein phosphatases (PPase) genes [Cell signaling, gene expression and mitosis but also CWI are cellular processes regulated by phosphorylation/dephosphorylation . Based oe) genes . As defee) genes ,130,131.S. cerevisiae cells treated with caffeine, PUF5 and POP2 have the same genetic pathway, leading to the conclusion that the CWI functions are mediated by Puf5 or Pop2-mediated gene repression mechanisms [Caffeine, as a CWI pathway activator, was used to show that Puf5 has a role in response to DNA replication stress and does not involve Pop2. Puf5 is a prototypical PUF protein, a family of RNA binding proteins conserved in eukaryotes, with roles in cell growth, division, differentiation, and development . In S. cchanisms .Kcs1 display defective vesicular endocytosis, showing a decrease in cell growth [kcs1Δ mutants are resistant to its lethal effects. Indeed, the lethal action of caffeine is suppressed in mutants that cannot synthesize inositol pyrophosphates because they physiologically antagonize the actions of Tel1 and Mec1 kinases [Inositol hexakisphosphate (IP6) is the most abundant inositol polyphosphate present in eukaryotes. IP6 is phosphorylated by IP6 kinases (IP6K-s) yielding inositol pyrophosphates, which are important signaling molecules in the eukaryotic cell . Yeast ll growth , sensitil growth , and abnl growth . Inosito kinases .S. cerevisiae to understand different molecular mechanism highly conserved in superior eukaryotes and to elucidate the way of action of compounds with potential as human drugs are presented below.Other examples of using the pleiotropic action of caffeine on S. cerevisiae proteotoxicity model of PD was employed to evaluate the role of caffeine in the aggregation of α-synuclein. On caffeine treatment, the toxicity of aggregates decreased, the intracellular oxidative stress was diminished, and the survival of the cell increased. It is supposed that caffeine alters the aggregation pathway of α-synuclein by introducing species with reduced proteotoxicity, leading to a decrease of the lag time and an increase in the apparent rate of fibrillation of α-synuclein. α-Synuclein has the ability to assume alternate aggregation pathways more than any other protein that apparently is misfolded in neurodegenerative disorders, because of its natively disordered structure. This effect apparently is heightened by the presence of caffeine, supporting the epidemiological studies that showed that coffee consumption is inversely related to the risk of onset of PD [The major component of Lewis Bodies ) is the natively disordered protein, α-synuclein . A S. ceet of PD ,140.S. cerevisiae cell by exposure to glucose, acting on a crossover point at the level of the phosphofructokinase/fructose-bisphosphatase cycle, increasing the ATP levels. Following glucose entry into the cell, caffeine reduces the concentration of intracellular cAMP in a dose-dependent manner, an effect that can be explained by the interference with catabolic inactivation of enzymes [Early studies also introduced caffeine as an activator of the cAMP-dependent protein kinase pathway, based on the in vitro potency of this compound to inhibit the mammalian cAMP phosphodiesterase . In yeas enzymes .S. cerevisiae defective in genes involved in the biosynthesis of mitochondrial phosphatidylglycerol and cardiolipin, pell and crd1, were analyzed in the presence of different cell wall perturbing agents. The mutants containing dysfunctional mitochondria revealed a modified sensitivity to metabolic inhibitors. The S. cerevisiae pell mutant showed increased sensitivity to the cell-wall perturbing agents such as caffeine, caspofungin, and hygromycin [Mitochondria play a fundamental role in eukaryotic cell physiology by integrating numerous death signals, being involved in the control of apoptosis. Mitochondrial genome integrity is essential for the viability of most species. Two mutants of gromycin .S. cerevisiae is an excellent model to study LS as it has conserved ageing pathways, and the study of new molecules’ effect on LS is greatly facilitated by yeast studies which yield significant information before proceeding to animal studies [The ageing biology is a new field that emerged since researchers have been attempting to extent the organisms’ lifespan (LS), and caloric restriction is a critical method used to understand the mechanism of LS. Because caloric restriction is usually accompanied by a reduction in food consumption over a long period of time, chemical food substitutes called caloric restriction mimetics have been the topic of intense research, and they can be used as starting materials in developing drugs that prevent or ameliorate the ageing-associated illnesses. Such compounds have shown their ability to extend the LS in different model organisms, e.g., rapamycin in mice  and yeas studies .Physical exercise, caloric restriction, and consumption of moderate amounts of substances such as selenium, zinc, omega 3 unsaturated fatty acids, vitamins E and C, antioxidants, caffeine, or alcohol were proposed as factors essential to extend the human LS or to reduce age-associated diseases. Often, these studies show only correlative (not causative) effects between a compound and longevity. The conservation of most ageing pathways in yeast and their facile genetic manipulation represents a premise to distinguish between the correlative and causative effects of nutrition on ageing .S. cerevisiae for identifying conserved genetic and pharmacological interventions [tor1Δ mutant cells. Studies correlating caffeine, TOR pathway, and LS have since been done on budding yeast [The extent of the LS was studied on ventions . The TORventions . Among tng yeast ,87 and fng yeast , inverteng yeast ,153 and ng yeast ,154.Caffeine treatment of yeast cells releases Rim15 from TORC1-Sch9-mediated inhibition and as a result, it increases LS. Therefore, it is highly probable that an analogous mTORC1/S6K/LATS kinase cascade also has influence on longevity in higher eukaryotes, including humans ,156. It S. cerevisiae was studied under two settings: in the stationary phase reached after glucose depletion and under severe caloric restriction. It was observed that cocoa polyphenol-rich extracts increased the chronological LS of S. cerevisiae during the stationary phase in a dose-dependent manner and also extended yeast LS under severe caloric restriction conditions. The cocoa extracts increased the lifespan of wild type cells and also of the sod2Δ cells, proving that the mechanism is Mn-SOD2-independent. Nevertheless, this effect was detected only for the polyphenol-rich cocoa extract and not for its individual components, including caffeine [The effect of a polyphenol-rich extract from cocoa on the chronological LS of caffeine .S. cerevisiae under conditions of caloric restriction. In contrast with other studies, caffeine has been claimed to increase the LS of yeast [Caffeine  was evaluated as a LS extender in of yeast , while oof yeast . Haploidof yeast .Pathologic endogenous DNA double-strand breaks (EDSB) can occur spontaneously even without exposure to radiation or DNA damaging agents . EDSB caS. cerevisiae, contributed to elucidating molecular mechanisms involved in biologic processes of general concern, such as DNA repair mechanisms, cancer, or aging. Using various approaches and setting multiple targets, the studies on caffeine–S. cerevisiae interaction generated outputs which could be extrapolated to higher organisms. In spite of the pleiotropic effects of caffeine, there is one mechanism universally accepted, i.e., the inhibitory effect on PI3 kinases, including the core kinases from the TOR complexes. However, since many of the puzzle pieces are still missing, it is no doubt that the duo caffeine–S. cerevisiae has not yet reached its full potential in opening doors to new knowledge.Caffeine, one of the most consumed and widely accepted neurostimulants, is also a powerful agent used in life science research. Due to its pleiotropic effects , caffein


We analyzed the associations between area-level socioeconomic status (SES) and prehospital delay in acute ischemic stroke (AIS) patients by degree of urbanization with the use of an ecological framework. The participants were 13,637 patients over 18 years of age who experienced AIS from 2007 to 2012 and were admitted to any of the 29 hospitals in South Korea. Area-level SES was determined using 11 variables from the 2010 Korean census. The primary outcome was a prehospital delay (more than three hours from AIS onset time). Multilevel logistic regression was conducted to define the associations of individual- and area-level SES with prehospital delay after adjusting for confounders, which includes the use of emergency medical services (EMS) and individual SES. After adjusting for covariates, it was found that the area-level SES and urbanization were not associated with prehospital delay and EMS use was beneficial in both urban and rural areas. However, after stratification by urbanization, low area-level SES was significantly associated with a prehospital delay in urban areas (adjusted odds ratio (aOR) 1.24, 95% confidence interval (CI) 1.04–1.47) but not in rural areas . Therefore, we posit that area-level SES in urban areas might be a significant barrier to improving prehospital delay in AIS patients. Stroke remains one of the most devastating of all neurological conditions. Globally, it accounts for approximately 5.5 million deaths annually, with 44 million disability-adjusted life years lost. As a disease of aging, the prevalence of stroke is expected to increase significantly around the world . The pasAmong the many risk factors of prehospital delay, some researchers believe that low socioeconomic status is one of the causes ,9. MoreoThus, we used an ecological framework to compare the associations between area-level SES and prehospital delay among AIS patients based on the degree of urbanization. Data from different regions and institutions should be sufficiently collected to achieve urbanization stratification. However, such data is challenging to collect and manage. Moreover, indicators for measuring the level of deprivation in each area should be applied to reveal the effect of SES. In this study, the authors attempted stratification by using multicenter data, targeting the whole country. By using a valid deprivation index, we could measure the effect of SES reasonably. Moreover, as far as we know, this is the first study to apply a multilevel approach to access prehospital delay.There are 17 administrative divisions in South Korea: nine provinces , six metropolises, one “special city,” and one “special self-governing city.” These divisions are subdivided into several smaller entities, including cities (si), counties (gun), and districts (gu). Provinces, the highest administrative category in Korea, are divided into counties. In the case of an urbanized area with a population of over 50,000 among counties, it can be independent as a city. Additionally, among them, cities with a population of over 500,000 are divided into districts. Therefore, in this study, districts of large cities, undivided cities, and other counties were applied at the same level of regional units. The determinant for urban area, based on the 2010 population census, is districts within a city having a population greater than 500,000. Finally, the country was divided into 248 regional units, of which 107 units belong to the urban area category and 141 units belong to the rural area category. For each unit, the area-level SES index, which will be described later in this section (2.3.1. Area-Level Socioeconomic Index), was measured.South Korean emergency medical services (EMS) are provided by the National Fire Agency through a Basic Life Support (BLS) single-tier system, a system that dispatches the same level of EMS regardless of the patient’s condition. Thus, the administration of intravenous thrombolysis by an EMS provider before hospital arrival is illegal .The Cardiovascular Disease Surveillance (CAVAS) project is a nationwide prospective stroke registry sponsored by the Korean Centers for Disease Control . PatientThe study included all adult patients (aged 18 years and above) to whom the International Classification of Disease-10th Revision code I63 or I64 was assigned on emergency department discharge between 1 November 2007 and 31 December 2012. Patients whose stroke onset or hospital arrival time data were missing were excluded, as were those who arrived 24 h after the onset of symptoms. Those without data on EMS use, address, age, sex, or individual-level SES were excluded, as well .Area-level SES was based on a deprivation index created by sampling 10% of the data from the 2010 Korean population census, recorded at the district level by the Korean National Statistics Office . Most deAll the regions were stratified into urban and rural at the provincial administrative level. If the province where the patient belonged had more than 500,000 people according to the 2010 census population, it became an urban area. In the urban areas, there were 47 (36.2%) districts belonging to the affluent category, 39 (36.4%) middle, and 21 (19.6%) deprived. In the rural areas, there were 20, 22, and 99  districts, respectively. The patients’ characteristics related with urbanization are shown in The CAVAS-derived education level and occupation status data were used to denote individual SES. The education level was divided into high school graduates and non-high school graduates. Occupations were then classified using the 10 major types of the Korean Standard Classification of Occupations based on the International Standard Classification of Occupations . We diviWe retrieved age, sex, and clinical information, times of AIS onset and hospital arrival, EMS use, and interhospital transfer status from the CAVAS registry. We then divided the patients into three age groups . The onset time was divided into day (6 a.m. to 6 p.m.) and night (6 p.m. to 6 a.m.) .The primary outcome was a prehospital delay (more than three hours from AIS onset time). The AIS onset time corresponds to the time at which the patient or caregiver first noticed symptoms or to the last known normal time if symptoms were not witnessed. According to the treatment guidelines published in 2018 , IV altep = 0.0014), and there were differences in the variables of various prehospital stages depending on urbanization. Statistical significance was decided with 95% confidence intervals (CIs). The multilevel logistic regression models were generated using the GLIMMIX procedure of SAS ver. 9.4 .Summary statistics were presented as median with interquartile range for non-normal distributed variables and as frequency with proportion for categorical variables. Differences in prehospital time intervals by urbanization status or area-level SES were assessed using the Wilcoxon rank-sum test and Kruskal–Wallis test. We used the chi-squared test to compare the groups in terms of area-level SES. Subsequently, we evaluated hierarchically to determine the effect of individual SES and area-level SES on prehospital delay using multilevel multivariable logistic regression. A random intercept-only model  was used to analyze the area-level variance in SES, and intraclass correlations were calculated. The associations of individual SES parameters with prehospital delay were assessed and specified as Model 1. Model 2 was evaluated by adding area-level SES variables to Model 1. Furthermore, the characteristics of patients significantly related to prehospital delay in univariate analysis were included in Model 3. Model 3 was further evaluated by stratification according to urbanization. Stratification by urbanization was carried out for two reasons: the interaction terms of area-level SES and urbanization were significant when the fitness was tested .Out of 30,552 adult AIS patients, 13,637 were qualified to be included in this study . Of thisOf all participants, 60.3% had not completed high school, 55.6% were unemployed, and 26.8% were manual workers. The affluent group included more high school graduates and non-manual workers. The proportion of Medicaid patients was highest in the deprived group .p < 0.001) (p = 0.04).Among all the patients, 59.4% experienced a prehospital delay. The median time from AIS onset to hospital arrival was 4.3 h (interquartile range (IQR) 1.7–10.4), 4.0 h (IQR 1.4–10.0), and 4.6 h (IQR 2.0–10.8) for the affluent, middle, and deprived groups, respectively (< 0.001) . Of the p < 0.001). This means that area-level SES was significantly associated with prehospital delay. Considering the difference in the variance of random effects, individual-level SES explained 3.0% of the variance in prehospital delay (basic model and Model 1), and area-level SES accounted for an additional 6.8% of the variance in prehospital delay (Model 1 and Model 2).The odds for a prehospital delay in the deprived group were 1.28 times higher than the odds in the affluent group in the unadjusted model (odds ratio (OR): 1.28; 95% CI: 1.10–1.48). However, the difference was not significant after adjusting for all covariates (adjusted OR (aOR): 1.03; 95% CI: 0.89–1.20). A higher education level predicted a low probability of prehospital delay in the unadjusted model  but not in the final model  .In urban areas, the odds for a prehospital delay were 1.24 times higher in the deprived group  than those in the affluent group, and the odds among economically inactive patients were 1.35 times higher than those among non-manual workers . In rural areas, we found no significant association between area-level SES and prehospital delay. The “other” occupational group was significantly more likely to experience prehospital delay compared to non-manual workers  in rural areas . EMS useIn this study, we examined whether area-level SES was associated with prehospital delay for AIS patients according to their degree of urbanization. Patients from deprived areas experienced approximately a 36 min longer delay compared to those from affluent areas. The association between area-level SES and prehospital delay differed according to the degree of urbanization; those classified as deprived in the urban areas faced a higher risk of delay than those in other area-level SES groups. However, there was no difference in the risk of prehospital delay between the deprived and the affluent in rural areas. A unique feature of this study is its use of a deprivation index that determines the SES of residential areas based on various factors. Given that SES is a complex metric that comprises multiple social factors, we tried to collect various types of information that reflect individual- and area-level SES to obtain an accurate representation of the SES level. Through the multilevel analysis, which included both the individual- and area-level SES, we aimed to determine the amount of variance in prehospital delay explained by both these types of SES.Prehospital delay is one of the major concerns of AIS treatment, and studies of cardiovascular care have reported some inequality in service and treatment provision for those in low-SES groups . In AIS,As shown in Interestingly, among patients who used EMS, the time of the call from the onset of symptoms was significantly longer in the deprived group than in the affluent group ; howeverFor patients who were transferred from another hospital, even if the time from symptom onset to arrival at the first hospital was shorter for patients in rural areas, the time from symptom onset to arrival at the final hospital was significantly longer for patients in rural areas compared to that for those in urban areas . For patEMS use and interhospital transfer are crucial for reducing prehospital delays ,26,27,28The strengths of this study were that it used a nationwide dataset, included patients regardless of EMS usage, was stratified by urbanization, and focused on the gap of SES as well the absolute level of SES.Like most studies, this study has its limitations. First, we excluded 7823 patients because of missing data, which might have introduced sampling bias. However, an analysis of the home addresses of the 1802 patients whose addresses were recorded but whose other data were missing revealed similar proportions in different area-level SES groups ; middle group, 606 (33.6%); deprived group, 589 (32.7%)). Therefore, the exclusion of these patients did not significantly affect the results of this study. Finally, given the possible differences in social structure, education, and health services, the results may differ in other countries. Thus, further research is required to confirm this.Area-level SES is not associated with a prehospital delay in AIS patients after adjusting for covariates. However, residing in a deprived urban area may increase prehospital delay, although this does not appear to be the case in rural areas. Area-level SES in urban areas might be a significant barrier to reducing prehospital delay in AIS patients. Finally, EMS use plays a crucial role in reducing prehospital delay, independent of urbanization status and area-level SES. EMS personnel must, therefore, consider potential preconceived ideas or biases for low-SES patients.


Here we report a case of pancreatic fistula due to obstruction of the jejunal loop due to compression of the jejunal loop by the superior mesenteric vessels.Pancreatic fistula is a common complication after pancreaticoduodenectomy, which could be caused by: soft pancreatic tissue, pancreatic duct diameter < 3 mm and body mass index ≥25 kg/mA 68-year-old man was admitted to our ward due to intermittent epigastric distension and pain. After various examinations and treatments, he was diagnosed with middle bile duct cancer. Pancreaticoduodenectomy was performed, and pancreaticojejunostomy and hepaticojejunostomy were completed by lifting the jejunal loop from behind the superior mesenteric vessels to the upper region of the colon. On postoperative day 9, the patient developed acute diffuse peritonitis, and on postoperative day 10, the patient underwent a second exploratory laparotomy, during which it was confirmed that the pancreatic fistula was caused by obstruction of the jejunal loop due to compression of the jejunal loop by the superior mesenteric vessels, then the patient recovered and was discharged alive after retrograde drainage in the jejunum.The superior mesenteric vessels after pancreaticoduodenal surgery can compress the jejunal loop and cause obstruction leading to serious complications, and it is recommended that general surgeons should avoid lifting the jejunal loop from the posterior aspect of the superior mesenteric vessels to complete the anastomosis. Pancreatoduodenectomy (PD) is advocated for treating most malignant and benign neoplasms of the pancreatic head and periampullary region . PD proc2, and have been proven to be independent risk factors for POPF [Several factors contribute to the development of a POPF, and every pancreatic surgeon must be aware of these factors. Patient-related factors can influence POPF development, including soft pancreatic tissue, a small pancreatic duct diameter of < 3 mm, and a body mass index ≥25 kg/mfor POPF –10. MoreIn this report, we present one case of Grade-C POPF after pancreaticojejunostomy due to the completion of anastomosis in the upper mesenteric vessels behind the jejunum loop obstruction extraction, along with a review of the literature.A 68-year-old man was admitted to our ward with intermittent epigastric distention for more than 1 month. The patient underwent surgery for penile cancer 21 years ago. The physical examination was unremarkable, with the exception of slight tenderness in the right upper abdomen. The patient’s height was 175 cm, weight was 70 kg, and body mass index (BMI) was 22.86. The initial laboratory results showed: total bilirubin (TBIL) 17.9 umol/L, direct bilirubin (DBIL) 9.3 umol/L, alanine aminotransferase (ALT) 526.5 U/L, aspartic aminotransferase (AST) 302.1 U/L, alkaline phosphatase 1756.6 U/L, glutamyl transpeptidase (GGT) 2240.6 U/L, and carbohydrate 19–9 (CA19–9) 26.11 u/ml. The initial leucocyte count, platelet count, renal function, coagulation profile, and other electrolytes were normal. Upper abdominal magnetic resonance imaging (MRI) showed local wall thickening enhanced in the upper pancreatic segment of the common bile duct and lumen stenosis, mostly considering choledochal carcinoma, with upper bile duct dilation and without evidence of invasion of the superior mesenteric vessels and metastasis  showed a small amount of fluid in the abdominal and pelvic cavities, a dilated bowel, and effusion in the upper abdominal bowel cavity, which was considered to be an obstruction. The patient was admitted to the intensive care unit and received conservative treatment, including fasting, gastrointestinal decompression, anti-infective treatment, proton pump inhibitors, somatostatin, and analgesia. On POD 10, the heart rate was 170 beats/minute, BP was 108/59 mmHg, leucocyte count was 13.15 × 109 cells/L, and drainage amylase level was 145.5 U/L. Reexamination using total abdominal CT suggested bilateral subdiaphragmatic free gas, abdominal and pelvic effusion, anastomotic leakage  1, the endotracheal intubation was removed and gastrointestinal function was restored. A liquid diet was then given on POD 5 and a semi-liquid diet was given on POD 7. During this process, the vital signs and drainage amylase test results of the patient were normal. On POD 9, the patient suddenly experienced severe epigastric pain accompanied by nausea and vomiting. Physical examination revealed tension in the abdominal muscles, tenderness throughout the abdomen, and rebound pain. The patient’s heart rate was 130 beats/minute, blood pressure (BP) was 106/67 mmHg, leucocyte count was 8.25 × 10Considering anastomotic leakage, acute diffuse peritonitis, and septic shock, the patient underwent a second exploratory laparotomy. During the operation, the superior mesenteric vessels were found to be compressing the jejunum, resulting in obvious dilation of the jejunum loop in the upper colon region, and the jejunum in the lower colon region was empty. In front of the pancreatojejunostomy location, there was a leak with a diameter of about 5 mm, and a large amount of fluid surrounded the liver and spleen. A retrograde gastric tube was implanted in the jejunum from the proximal end of the Braun anastomosis to near the chojejunostomy location, a nutrition tube was implanted in the distal jejunum under the Braun anastomosis, several silicone drainage tubes were placed around the liver, spleen, and pelvis, and the operation was completed Fig. b.Fig. 4On the post-second operative day (PSOD) 1, the patient’s temperature was 37.9 °C, pulse rate was 90 times/minute, BP was 120/70 mmHg, and the endotracheal tube was removed after the patient awoke. Following that, enteral nutrition was performed, peritoneal lavage and drainage were continued, and anti-infective agents, proton pump inhibitors, somatostatin, and other treatments were continued. On PSOD 32, the patient’s pancreatic fistula was healed and the abdominal drainage tube was removed. The patient was discharged on PSOD 35. On PSOD 45, the nutrition tube was removed and the gastric tube in the jejunum loop was removed, indicating full recovery of the patient.Thanks to advances in surgical techniques and perioperative management, the current mortality rate after PD procedures has decreased to less than 5% in high-volume centers , 12, but2 [It is known that several factors contribute to the development of a POPF. There are technical and perioperative factors, on the one hand, and patient-related factors, on the other hand . In 20132 . Other l2 , 19, and2 .A recent meta-analysis  comparinIn this case, we retrospectively found that the occurrence of pancreatic leakage was due to technical defects. The pancreaticojejunostomy was completed following the lifting of the jejunum loop behind the superior mesenteric vessels, the superior mesenteric vessels compressed the loop, causing obstruction of the loop, and the pressure in the intestinal cavity of the loop was increased due to the accumulation of digestive fluid, eventually leading to rupture of the pancreaticojejunostomy. Fortunately, laparotomy was performed actively and a retrograde gastric tube was implanted in the jejunum from the proximal end of the Braun anastomosis to near the chojejunostomy location, playing a positive role in the decompression of the jejunal loop. A nutritional tube was implanted into the distal jejunum to address the patient’s enteral nutrition problems. Eventually, the patient recovered and has survived to this day.In pancreaticoduodenectomy, lifting the distal jejunum behind the superior mesenteric vessels to complete the pancreaticojejunostomy and choledochojejunostomy could lead to possible risk of jejunal loop obstruction due to compression of the superior mesenteric vessels and may also cause serious consequences such as pancreaticojejunostomy rupture or choledochojejunostomy rupture, therefore, surgeons should avoid lifting the jejunal loop behind the superior mesenteric vessels. Grade C POPF after pancreaticojejunostomy is a serious challenge for pancreatic surgeons. Various factors that contribute to POPF development have been reported in the previous literature, but there is no report that compression of the superior mesenteric vessels on the loop of the jejunum leads to POPF. This case may serve as a warning to pancreatic surgeons.


Relatively sweet vegetables like carrot and sweetcorn were used more often than bitter vegetables such as broccoli and spinach. The described texture of most  wet ‘spoonable’ products was of the lowest complexity . CACFs would adequately expose children to cow’s milk and wheat but not to other common food allergens . If children’s diets include CACFs, non-commercial meals must be offered as well in order to meet nutritional guidelines related to the introduction of common food allergens, diversity of flavours, and complex textures for infants and toddlers.Optimal nutrition in early childhood fosters growth and development whilst preventing morbidity and mortality in later life. There is little research in New Zealand on commercially available complementary foods (CACFs). This cross-sectional study of the nutritional aspects and packaging of CACFs used data collected in four major supermarket chains in New Zealand in 2019 (Nutritrack). Of the 197 CACFs analysed, 43 (21.8%) were inappropriately recommended for consumption by children four months of age or older, 10 (5.1%) had added salt, and 67 (34.0%) contained free sugars. The majority ( Nutrition in the first 1000 days of life is critical for growth and development, lays the foundation for eating behaviour, determines food preferences, and influences health outcomes in later life ,3,4,5,6.The foods chosen for children by care providers are dependent on complex social, economic, cultural, and political determinants of the food environment ,4,5,6,7.Global Strategy on Infant and Young Child Feeding encourages World Health Organization (WHO) member states to create national policies and strategies for infant feeding  children were packaged in squeezable pouches.In 2019, there were 197 discrete CACFs in New Zealand supermarkets, retailing under twelve different brand names. There were 153 (77.7%) wet ‘spoonable’ CACFs and 44 dry products. The most abundant category of CACFs was main meals  . All fisn = 161, 81.7%) of all CACFs were recommended for children less than 12 months of age [Stages 1–4]. As shown in The majority  of these CACFs were of the lowest textural complexity . Analysis as per product description showed that textures became more complex along the age-gradient for all vegetable-based, meat-based, and poultry-based main meals . HoweverTen (5.1%) CACFs were found to contain an added salt and these were from the following food sub-groups: two each of breakfast cereals, meat-based meals, and vegetable-based meals; and four rusks. Five (2.5%) of the CACFs were recommended for Stage 5 children aged more than 12 months old and five were for children six to twelve months old (Stages 2–4). The amount of sodium in all products recommended for children less than 12 months of age was below the regulatory thresholds.n = 67, 34.0%) of CACFs available for purchase contained free sugars. These were in the form of sugars , maltodextrin, fruit juices , dextrose, fruit juice concentrates , and glucose.More than a third , mostly in the form of cereals in breakfast meals and flour in main meals. Hen’s egg was present in four (2.0%) main meals as a constituent of pasta. Soy was present as flour in four (2.0%) products. Fish was present in only one (0.5%) product. Peanut, tree-nuts, and crustacean shellfish were absent as ingredients in all CACFs available for purchase in New Zealand supermarkets.Cow’s milk was used in 67 (34.0%) of the CACFs in the form of whole milk, yoghurt, and cheeses. Wheat was present in almost a quarter of CACFs , followed by bananas , pears , mangos , and blueberries . There were 24 discrete types of vegetables used in CACFs, of which carrots , onions , sweet potatoes , tomatoes , and pumpkin  were used more frequently. Relatively sweet vegetables like carrots, sweet potatoes, and sweetcorn were used more predominantly in CACFs than bitter vegetables such as spinach, broccoli, and cauliflower (There were 175 (88.8%) CACFs that contained at least one fruit and or vegetable. Of these, 93 (47.2%) contained only fruits, 61 (31.0%) contained only vegetables, and 21 (10.7%) contained both fruits and vegetables. There were 21 discrete types of fruits present in CACFs of which the most commonly used were apples (liflower .n = 136) of CACFs would impart an overall sweet flavour as they contained sweet flavoured ingredients. All ready-to-eat breakfast meals (n = 22), desserts (n = 21), fruit-based meals (n = 33), and sweet snacks (n = 29) contained sweet ingredients. Sixty percent of rusks (n = 3) and dry cereals (n = 6), 63.2% of vegetable-based meals, and 76.9% of fruit and vegetable-based meals contained sweet ingredients. All of the fish-based (n = 1), meat-based (n = 30), and poultry-based (n = 14) meals had ingredients with savoury flavours. There were 17 products (8.7%) that contained bitter vegetables and they may impart an overall bitter flavour in the product. However, these same 17 products contained one or more sweet vegetables as an ingredient too.Sixty-nine percent  of Stage 1 [4m+] CACFs  CACFs . Further+] CACFs .n = 103, 52.3%). Most  CACFs packaged in squeezable pouches had little textural complexity and were described as puréed, creamy smooth, smooth, or mashed. Such CACFs can be suckled directly from the packages, impeding oral development that is usually stimulated from chewing complex textures [The most prevalent form of packaging in New Zealand was the squeezable pouch  which was not in line with the Food and Nutrition Guidelines. Our results were congruent with findings of studies conducted in the United Kingdom, the United States, and Canada, which have found that only a few CACFs in those markets contained added salts [A small proportion of CACFs contained added salt  of CACFs contained free sugars despite the Food and Nutrition Guidelines recommending that sugar, honey and other sweeteners not be added to complementary foods [A significant proportion n = 67, 3.0% of CAry foods ,30,50,51ry foods .n = 136) of CACFs contained ingredients that would impart an overall sweet flavour. While 17 (8.7%) different products did contain a bitter vegetable, they also had sweet vegetables like carrots, parsnip, sweetcorn, sweet potatoes, and pumpkin as ingredients. The bitterness in foods can be damped by the addition of sweet and salty flavours [Food and Nutrition Guidelines recommend that children should be introduced to diverse flavours one at a time and repeatedly to enable learning and acceptance [n = 36, 18.3%) was noted in our study too. Manufacturers should provide more CACFs that would enable learning of bitter flavours by reducing reliance on fruit-based ingredients and sweet vegetables.This analysis showed that 69.0%  contained cow’s milk and almost a quarter  contained wheat, children fed a variety of CACFs would be adequately exposed to these two allergens in their first year of life. However, children would need to be fed non-commercial meals containing soy, cooked hen’s egg, fish, crustacean shellfish, peanut, and other tree nuts as CACFs are unlikely to expose them to these food allergens.The risk of developing food-induced allergies to common food allergens is reduced when these foods are introduced to the general low-risk population in their first year of life as compared to when delayed ,20. SincThe major strength of this cross-sectional study is the use of data collected by means of robust and reliable methods which represented 75.0% of the commercial foods available for purchase. A limitation was that CACFs marketed in specialty shops, ethnic outlets, and online were not included, although this is likely to be a very small proportion of total CACFs available on the market. Secondly, the nutritional information panel was utilized to extract the macronutrient content rather than the chemical analysis of the product itself. Actual values of nutrients may vary as manufacturers calculate the nutrient content from individual raw ingredients . HoweverAlthough CACFs are not the only source of nourishment for children, they do comprise a significant proportion of the diet of children aged six to twenty-four months old in New Zealand. This first descriptive analysis of CACFs in New Zealand has found that most products meet the current recommendations and regulations for salt and sodium but were largely sweet (due to free sugars and relatively sweet ingredients), lacked variety in the vegetables included, and did not contain common food allergens. A fifth of CACFs (21.8%) may encourage the displacement of nutritionally adequate breastmilk and substitutes as they were recommended as appropriate for children from four months of age. Children fed CACFs will also need to be fed nutritionally adequate homemade foods in order to introduce common food allergens and diversity of flavours. Our findings will have implications for all stakeholders. Practitioners must be mindful of the advice they provide regarding complementary feeding. Healthcare organizations must ensure that their endorsements do not contradict national guidelines and regulations. Carers must include non-commercial sources of complementary foods in the diets of children. The authors recommend that dietary guidelines and regulations be updated to reflect current global recommendations so that children are provided with the most appropriate, adequate, acceptable, and safe complementary foods.


CTNNB, PFDN5 and EIF3M, selected from RNA sequencing data, were the most suitable reference genes. The present study provides a detailed analysis of the expression stability of reference genes for the study of gene expression profiling during the transition process from BAT to WAT.Brown adipose tissue (BAT) plays important roles in unique non-shivering thermogenesis. It is necessary to select reference genes during the transition process from brown (BAT) to white adipose tissue (WAT) for Quantitative PCR (qPCR) analysis. In this study, PGC-1α and GPAT4. It was found that traditional reference genes, such as GAPDH, RPLP0, HPRT1, and PPIA were not suitable for target gene normalization. In contrast, CTNNB, PFDN5, and EIF3M, selected from RNA sequencing data, showed the least variation and were recommended as the best reference genes during the transformation from BAT to WAT.Brown adipose tissues have unique non-shivering thermogenesis functions, can be found in newborn ruminate animals, and then are gradually replaced by white adipose tissues in adulthood. For the purpose of exploring the intrinsic mechanism underlying the conversion process from brown (BAT) to white adipose tissue (WAT), it is necessary to utilize Quantitative PCR (qPCR) to study gene expression profiling. In this study, we identified reference genes that were consistently expressed during the transformation from goat BAT to WAT using RNA-seq data. Then, twelve genes were evaluated as candidate reference genes for qPCR in goat perirenal adipose tissue using three tools . In addition, the selected reference genes were used to normalize the gene expression of  It is well known that three types of adipose tissues exist in mammals, each with their own unique marker genes, which can also be differentiated from their morphology ,2. WhiteQuantitative PCR (qPCR) technology can quickly and accurately quantify the gene expression at transcriptional levels. The qPCR method has been reported to be more specific and powerful to determine the gene expression at different developmental stages or under different conditions ,10. HowePGC-1α and GPAT4 were taken as the target genes to correct the stability of candidate reference genes.It has been shown that brown adipose tissue is mainly found in the intrascapular, perirenal, and pericardial areas of newborns. With an increase in age, it is gradually replaced by white adipose tissue and adult animals have very little brown adipose tissue ,14. In oAll research involving animals was conducted according to the approved protocols of the Institutional Animal Care and Use Committee at the College of Animal Science and Technology, Sichuan Agricultural University, Sichuan, China, under permit No. DKY-B20176925.In the present study, Chuanzhong black goats were raised at the breeding center of the Sichuan Agricultural University, Ya’an, China . The annual average temperature and humidity in Ya’an are about 14 °C and 52%, respectively. The Chuanzhong black goat is one of the most important breeds for meat production in the southwest of China. A detailed description of the experimental design and sample collection was provided in our previous study . The perTotal RNA was extracted using RNAiso plus . RNA concentration was measured using a NanoDrop 2000 spectrophotometer . The RNA purity was distinguished by the ratio of OD260/280, and RNA quality was evaluated by RIN (RNA Integrity Number) value, which was calculated using the Agilent 2100 Bioanalyzer System . Each sample was reverse transcribed with 1 μg total RNA using the Reverse Transcription Kit .To identify genes whose expression had a low variation during the transformation from goat BAT to WAT, we retrieved 12 RNA-seq datasets previously published in relation to the transformation from BAT to WAT . The rawThe primer pairs of reference genes were designed by Primer Premier 5.0 software. The information of primers was shown in delta-Ct . Moreover, the data processing on BestKeeper was based on raw Ct-values from qPCR.The gene expression was detected by Bio-Rad CFX 96 . Then, the data were imported into geNorm, NormFinder, and BestKeeper, as required by each program and according to their manuals. Thus, the delta-Ct method was used to transfer Ct values to linear-scale expression quantities. The delta-Ct formula is as follows: Q = E r) value.The highest M value of the reference gene in geNorm is considered to be the most stable gene. A pairwise variation V value below 0.15 indicates that the inclusion of an additional reference gene is not needed. The algorithm of NormFinder estimates the overall expression variation and the variation between subgroups of the sample set. In addition, the lowest stability value of the reference gene is considered to be the most stable gene in the sample set investigated. In the BestKeeper tool, pairwise correlations and regression analysis are used as process approaches. The highest stability reference gene has the lowest values for the coefficient of variation (CV) and standard deviation (SD), but has the highest correlation coefficient  were selected for further validation. Among them, CTNNB1 and PFDN5 genes have the lowest coefficient of variation. In addition, we selected four reference genes  according to previous studies in white and brown adipose tissue [GAPDH, 18SrRNA, YWHAZ, and ACTB genes [Based on the FPKM value and the coefficient of variation, we obtained a total of 54 candidate reference genes . KEGG , followed by RPL22, YWHAZ, ACTB, RPLP0, and HPRT1. The most variable reference genes were 18SrRNA, PPIA and GAPDH. Interestingly, three novel reference genes  that were selected from RNA-Seq data mostly expressed good stability. In the pairwise variation analysis, we found that the optimal number of reference genes for normalization was three, using 0.15 as a cutoff value  by comparing a specific gene with all of the other reference genes and the average pairwise variation was used to select the optimal number of reference genes. The reference gene with the smallest M value had the strongest stability. As shown in ff value B.PFDN5, EIF3M, CTNNB1, TBP, and ACTB were the most stable genes, with stability values less than 0.15. Meanwhile, YWHAZ, 18SrRNA, and RPL22 showed a relatively lower stability. However, RPLP0, HPRT1, GAPDH, and PPIA were the least stable reference genes.NormFinder calculates the intragroup and intergroup variations. As shown in r), standard deviation (SD) and coefficient of variation (CV). So, the two least stable reference genes in geNorm were excluded. Firstly, the candidate genes have an acceptable overall variation to be considered reference genes (SD < 1). Secondly, a coefficient of correlation (r) calculation was performed. Then, the stability was established based on the coefficient of correlation (r). The stability was mainly judged by the correlation coefficient (r), then by considering the standard deviation (SD). Thus, TBP, CTNNB1, and 18SrRNA were the most stable reference genes as r > 0.8, then PFDN5, EIF3M, and ACTB followed, as their r > 0.7. The correlation coefficient (r) value of YWHAZ, RPLP0, HPRT1, and RPL22 was less than 0.7, expressed by the bad stability in the results of BestKeeper  or the least stable reference genes  to normalize the same target genes. Using CTNNB1, TBP, PFDN5, and EIF3M for normalization, the results showed that PGC-1α and GPAT4 were expressed in perirenal adipose tissues with the highest level at D1, and significantly (p < 0.05) decreased at D30, then significantly (p < 0.01) reached the lowest level at Y1  plays important roles in unique non-shivering thermogenesis and is enriched with mitochondria . Althoug in mice .In addition, the three tools used in this study have different algorithms, so the stability sorting results of candidate reference genes need to be analyzed synthetically. In geNorm, in calculating the M value of a reference gene, the tool mainly focuses on the comparison between the reference genes, and only the differences between groups are taken into account . In NormACTB showed medium stability; however, ACTB was quoted as the least stable reference gene during 3T3-L1 adipocyte differentiation [GAPDH has been confirmed to have an unstable expression in mesenchymal stem cells during differentiation [PPIA is the most variable in all stages during 3T3-L1 adipocyte differentiation [PPIA is expressed with high stability among different goat tissues  [GAPDH and PPIA were both proven to be unstable candidate genes by geNorm and NormFinder, respectively. RPLP0 is the least stable gene in bovine (subcutaneous) and goat  adipose tissues in three physiological states  [In our results, ntiation . GAPDH hntiation  and showntiation . PPIA isntiation  and is vntiation , while intiation . Howeverd liver) . In thisand dry) , while iand dry) . HoweverPGC-1α and GPAT4 genes are the marker genes for brown adipose tissue [PGC-1α and GPAT4 genes were selected as target genes for normalization. PGC-1α is the first transcriptional regulatory factor for regulating brown adipose thermogenesis [PGC-1α and GPAT4 were, as expected, significantly downregulated in their expression from D1 to Y1, when the two target genes were normalized to CTNNB1, TBP, PFDN5, and EIF3M, the four best stability reference genes. However, severe variance appeared when they were normalized to RPLP0, GAPDH, HPRT1, and PPIA, the four least stable candidate genes. Hence, for the accuracy of qPCR, appropriate reference genes should be carefully selected, because the unstable reference genes will cause the misinterpretation of expression data.Previous studies have shown that e tissue . In addiogenesis ,35. GPATogenesis . In thisCTNNB1, TBP, PFDN5, and EIF3M were recommended as the most suitable reference genes in qPCR experiments during the transition process from brown to white adipose tissues.In summary, we found that


This study aimed to develop an explanatory model of sexual satisfaction in same-sex attracted individuals with a partner, based on personal and interpersonal variables. The participants were 410 men  and 410 women  who maintained a relationship with another person of the same sex. Internalized homophobia was considered as a personal variable, and as interpersonal variables, the dimensions of attachment (anxiety and avoidance), sexual functioning, dyadic adjustment, relationship satisfaction, the components of the Interpersonal Exchange Model of Sexual Satisfaction, the number of sexual costs and the number of sexual rewards were considered. The degree to which sexual satisfaction was related to these variables was examined separately, for both men and women, through multiple linear regression models within the framework of structural equation models. The results indicated that sexual satisfaction is associated in a negative sense with internalized homophobia, the number of sexual costs, anxiety, and avoidance, and in a positive sense with the remaining variables. Relational variables were more relevant in the explanation of sexual satisfaction. The clinical implications are discussed. Sexual health is not merely the absence of disease, but a state of physical, mental, and social well-being concerning an individual’s sexuality . On the Most studies focused on sexual satisfaction have focused their interest on the Caucasian/Anglo-Saxon heterosexual population ,9,10, anIn this organizational system, based on the Ecological theory ,16,17, tREW-CLCST), and (3) perceived equality of sexual costs (EQCST) and rewards (EQREW) between the members of the couple. Although not part of the model, the number of sexual rewards and the number of sexual costs were also considered as interpersonal variables.Personal and interpersonal variables have been shown to outweigh the social and ideological-cultural variables in the explanation of sexual satisfaction ,4. The pSexual minorities have become increasingly visible; as a result, the opportunity to examine different aspects of same-sex couple relationships has increased . Few stuWill personal and interpersonal variables associate (directly and/or indirectly) with sexual satisfaction?Will the explanatory models show gender differences?In addition to the hypothesis, it is worth mentioning that two research questions regarding the aforementioned variables were posed while doing this research:M] = 29.24, standard deviation [SD] = 9.84), and for women, between 18 and 58 . Most participants were Spanish (57.6%); the rest were Mexican (23.7%), Colombian (12%), Chilean (4.3%), Venezuelan (1.1%), Argentinian (0.5%), Dominican (0.2%), Ecuadorian (0.2%), Cuban (0.1%), Honduran (0.1), Paraguayan (0.1), and Peruvian (0.1). Exclusion criteria were: (1) Being a citizen of a non-Spanish-speaking country, (2) having Spanish as a non-native language, (3) being underage, (4) having a sexual orientation that is not towards people of the same sex, (5) discrepancy between biological sex and gender identity in the participant or their partner, (6) do not maintain a romantic relationship for at least three months, and (7) not having sexual activity with the partner at the time of the evaluation. A sample of 820 Spanish-speaking participants (410 men and 410 women) who had been in a romantic relationship for at least three months with a person of the same sex was obtained by nonprobabilistic convenience sampling. For men, the age range was between 18 and 66 , using Limesurvey® software  and controlling the IP address of the responses; to avoid automated responses, participants were asked to confirm their access to the survey by responding to a security question consisting of a simple randomized arithmetic operation. To distribute the questionnaire via Facebook®, a post was published on the Laboratory of Human Sexuality page and promoted to invite users of this social network to participate. Anonymity was guaranteed to all participants, as well as the confidentiality of their data, and their participation was voluntary. Before responding, participants were asked to read and accept an informed consent form, which described the purpose of the study and provided information on data confidentiality and privacy. The study was approved by the University of Granada Human Research Ethics Committee.Evaluation instruments were administered using both the traditional paper-and-pencil format and the online format. Neither method showed differences in the information collected on sexual behaviors . ParticiA sociodemographic questionnaire was used to collect information on sex, age, nationality, couple relationship, partner’s sex, length of relationship, cohabitation with partner, sexual relations with current partner, age of first sexual relation and type , and number of sexual partners.Exclusively heterosexual (1) to Exclusively homosexual (7); an eighth option was included to account for asexuality. Only participants who reported being exclusively attracted to people of the same sex were selected.The Kinsey Scale  was usedThe Spanish version of the Internalized Homonegativity (IH) scale , publishTotally disagree (1) to Totally agree (7). Higher scores indicate higher anxiety or avoidance. The instrument has shown good reliability in same-sex attracted population [The Spanish version of the Experiences in Close Relationship Short-Form (ECR-S) , adaptedpulation . In our Totally decreased (0) to Normal (4). Given that the possible range of scores was different for men and women, the mean score was used instead of the total score. To avoid overlapping between sexual functioning and the measure of sexual satisfaction, item 5  was excluded from the calculation of the mean. Higher scores indicate good sexual functioning. This questionnaire presents good reliability in the same-sex attracted population [The Spanish version of the Massachusetts General Hospital-Sexual Functioning Questionnaire (MGH-SFQ) , adaptedpulation . In the Always disagree to to Always agree to) or five response options (from Never to Every day), depending on the item. Higher scores indicate better adjustment. It has been shown to be reliable and valid for same-sex attracted population [The short Spanish version of the Dyadic Adjustment Scale (DAS) , adaptedpulation , and theNot at all rewarding (1) to Extremely rewarding (9). Item 2 evaluates the actual level of sexual rewards in comparison with the expected level thereof (CLREW) with anchors from Much less rewarding in comparison (1) to Much more rewarding in comparison (9). Item 3 evaluates the perceived level of reward in comparison to one’s couple’s perceived level of reward (EQREW) with anchors from My rewards are much higher (1) to My partner’s rewards are much higher (9). The three remaining items  are similar to the first three but focused on sexual costs (CST). The total balance between rewards and costs (REW-CST) is the difference between the score of item 1 and the score of item 4. The level of comparison between sexual rewards and costs (CLREW-CLCST) is calculated by subtracting the score of item 5 from the score of item 2. In both cases, possible scores range from −8 to +8, where higher scores represent more sexual rewards. In the present study, the components of equality of rewards (EQREW) and equality of costs (EQREW) were excluded because they have been shown to have little or no impact on sexually satisfied people [Very bad/Very good, Very unpleasant/Very pleasant, Very negative/Very positive, Very unsatisfying/Very satisfying, Very worthless/Very valuable. The GMREL is identical to the previous measure of satisfaction, but it measures satisfaction with one’s romantic relationship. Finally, the RCC is composed of 58 items, representing different sexual exchanges that can be valued as rewards, costs, both, or neither. The sum total of rewards indicates the number of sexual rewards and the sum total of costs represents the number of sexual costs. In the present study, the GMSEX obtained an ordinal alpha of 0.94 for men and 0.93 for women; concerning the GMREL, an ordinal alpha of 0.95 was obtained for both men and women.The Spanish version of the Interpersonal Exchange Model of Sexual Satisfaction Questionnaire (IEMSSQ) , publishd people ,54. The t-test for independent samples was used for each of the studied variables. Secondly, correlations between the evaluated variables were carried out. In this way it was possible to confirm whether the direction that the associations of the variables had with sexual satisfaction was positive or negative. The nationality variable was included in the aforementioned analysis to determine its relationship with the other variables. Subsequently, all the variables  were included in a structural equation model using RStudio software  [R2) has been shown without the fit indexes that are commonly evidenced. Therefore, for the development of explanatory or predictive models, the variance portion can be used as a unique criterion to show the explanatory, or predictive, capacity in models that are developed with SEM. Given the noncompliance of the multivariate normality, and designed for ordinal data [p values below 0.05 indicate statistical significance.Firstly, to analyze differences by sex, Student’s MA, USA)  and the MA, USA) . The expMA, USA)  and, thrMA, USA) . In thisMA, USA) . This puMA, USA) . In someMA, USA) ,60,61, Snal data , the ananal data . In all REW-CLCST, and number of sexual costs. No significant differences were found between men and women in regard to internalized homophobia  and number of sexual rewards . Men reported better sexual performance than women, whereas women reported more dyadic adjustment, sexual satisfaction, and relationship satisfaction, as well as more rewards than sexual costs (REW-CST) and a higher relative level of rewards than cost (CLREW-CLCST); compared to men, they reported lower anxiety, avoidance, and fewer sexual costs than men. However, both men and women reported good sexual and relationship satisfaction, as well as good dyadic adjustment and sexual functioning. Additionally, all participants presented low anxiety, avoidance, and internalized homophobia , number of sexual rewards, and number of sexual costs) observable variables were the independent ones.Given the sex-based differences in virtually all evaluated variables, independent models were created for men and women. Sexual satisfaction was the observable dependent variable in both models, whereas the rest of the personal  and interpersonal , relationship satisfaction (β = 0.61), REW-CST (β = 0.15), CLREW-CLCST (β = 0.13), and number of sexual rewards (β = 0.08) were directly and positively associated with sexual satisfaction. Additionally, the model included variables indirectly related to sexual satisfaction, as well as mediating variable. It was thus made apparent that relationship satisfaction acts as a mediating variable in the association between sexual satisfaction and internalized homophobia (β = −0.13), avoidance (β = −0.14), dyadic adjustment (β = 0.36), and number of sexual costs (β = −0.23); except for number of sexual costs, all associations were positive. In turn, internalized homophobia acted as a mediating variable between anxiety (β = 0.25) and relationship satisfaction , relationship satisfaction (β = 0.53), CLREW-CLCST (β = 0.23), number of sexual rewards (β = 0.11), and number of sexual costs (β = −0.06); in all cases, the association was positive, except for number of sexual costs. Number of sexual costs was found to act as a mediating variable between sexual satisfaction and internalized homophobia (β = 0.06), anxiety (β = 0.27), and avoidance (β = 0.18), all of them positively associated with CST. Relationship satisfaction was positively affected by dyadic adjustment (β = 0.53) and REW-CST (β = 0.18), acting as a mediating variable between these two variables and sexual satisfaction  reported by the study participants and the lack of differences by sex could be due to a secure attachment style  or perceIn terms of interpersonal variables, in general, both men and women obtained low anxiety and avoidance scores, which suggests secure attachment. In the case of women, our results are comparable to those reported by Guzmán-González et al. . AccordiOn the other hand, men obtained better sexual functioning scores than women, as has been the case in studies conducted with heterosexual populations ,73,74,75Concerning the sexual satisfaction explanatory models, few variables were found to be directly associated with sexual satisfaction. For men and women, these variables were sexual functioning, relationship satisfaction, comparison of sexual rewards and costs, and number of sexual rewards; specifically for men, sexual satisfaction was related to the balance between rewards/costs, and for women, to the number of sexual costs (research question 1). As hypothesized, all the variables were positively associated with sexual satisfaction, except for number of sexual costs, whose association was negative. These associations are consistent with previous research focused on same-sex attracted individuals ,16,32,78β = 0.61 in men and β = 0.53 in women) but also a mediating role between other variables and sexual satisfaction (research question 1). This is more relevant in the case of men because all the variables not directly associated with sexual satisfaction were instead associated with relationship satisfaction, that is, internalized homophobia, avoidance, dyadic adjustment, and number of sexual costs (research questions 1 and 2); in the case of women, relationship satisfaction was associated with dyadic adjustment and balance between sexual rewards and costs (research questions 1 and 2). As could be expected, the variable of dyadic adjustment (β = 0.36 in men and β = 0.53 in women) had the strongest influence on relationship satisfaction. Dyadic adjustment and relationship satisfaction refer to the quality of the romantic relationship; therefore, the close positive relationship between the two variables is logical and has been confirmed in other studies on same-sex attracted population [In both models, relationship satisfaction was found to have not only the highest weight in explaining sexual satisfaction . The fact that this mediating factor appears in the women’s model but not in the case of men could be due to several reasons. Concerning internalized homophobia, Cohen et al.  identifiIn men, anxiety acts indirectly through internalized homophobia and relationship satisfaction (research questions 1 and 2); the direction of these associations is also logical and as expected according to the hypothesis presented. The fact that anxiety has been associated with internalized homophobia in men but not in women could be due to several reasons. Attachment theory in adults  states tIn both models, relational and emotional variables (relationship satisfaction and dyadic adjustment) were found to explain most of sexual satisfaction, which makes them relevant variables when considering sexual satisfaction in the context of same-sex relationships. According to previous studies on same-sex attracted ,16,97 anThe present study had some limitations. In the first place, this investigation had a nonprobabilistic convenience sampling; therefore, the generalizability of the sample was limited and not representative. Secondly, due to the design of this research, the results did not establish causality effects since sexual satisfaction was assessed at a specific time. Thirdly, although people reported having a specific sexuality at the time of participating in this study, it is important to note that sexuality and relationship are fluid and dynamic. In the fourth place, only one personal variable was used; therefore, the study would need to be replicated using more personal variables . Fifth, all the associations showed unidirectional relationships, so it is suggested in future studies to incorporate reciprocal causal relationships in the associations that require it. Furthermore, sexual satisfaction was studied independently of the dyad formed by the couple; therefore, future research should focus on the sexual satisfaction of same-sex couples using actor-partner interdependence models (AIPM) to showcase the influence of one partner’s variables on the other’s ,103. In This study proposed explanatory models of sexual satisfaction for men and women with a same-sex romantic partner. The models included personal and interpersonal variables, in which the IEMSS components  and variIn sum, sexual satisfaction is regulated by both personal and interpersonal factors, with the relational aspects having the highest weight. These results are in line with studies showing that dyadic processes are a fundamental characteristic in the sexual satisfaction of people in a romantic relationship ,106.


The COVID-19 pandemic has had a severe impact worldwide. Developed countries, such as the United States, United Kingdom, Italy, and Spain, had their highly efficient medical infrastructure greatly stressed and suffered from high death tolls. Similarly, Bangladesh, a poverty-stricken South Asian country, is losing its battle against the pandemic, but mainly because of its incompetent healthcare system. The casualties are escalating and public sufferings are becoming unimaginable. On this backdrop, this perspective piece discusses the healthcare crisis in Bangladesh during the pandemic. This article also identifies three responsible issues for the country’s deteriorating health care: 1) poor governance and increased corruption, 2) inadequate healthcare facilities, and 3) weak public health communication. Rather, behind this mysterious façade, the reality tells a different tale. This pandemic reveals the incompetent health care of many less developed countries engulfed by intense corruption, and Bangladesh is one of them. As one of the world’s most densely populated countries  with 21.8% of people living below the poverty line,2 Bangladesh has a healthcare system that lacks reliability, responsiveness, and empathy, and that has already been proved inadequate to deliver proper health care to the public on many occasions.4 Moreover, medical facilities are concentrated in urban areas that create a healthcare divide depriving rural areas.5 Amid such a situation, the COVID-19 pandemic reveals many loopholes in the healthcare system that can be summarized under three themes: 1) poor governance and increased corruption, 2) inadequate healthcare facilities, and 3) weak public health communication.Surprisingly, whereas many developed countries, including the United States, the United Kingdom, Italy, and Spain, have suffered greatly during the COVID-19 pandemic even with their highly efficient healthcare systems, many less developed countries with inadequate healthcare systems are surviving the crisis more easily.6 Furthermore, a few of the government’s delayed decisions were found less effective that worsened the pandemic situation. For example, the government declared a general holiday for 10 days from March 23, 2020 to April 2, 2020 without restricting transportations and public movement. As a result, a crowd of more than 12 million people left Dhaka immediately after the notice that expedited the community transmission.7 Moreover, none of the two existing laws, Disaster Management Act 2012 and Infectious Disease  Act 2018, were brought into action properly to control such massive mass mobilizations and gatherings.6 In such a situation, many private medicals around the country were either unwilling or abstained from treating COVID-19 patients, and thus healthcare denial intensified.9 A survey found that 77.3% of patients receive healthcare facilities from private medicals.6 Consequently, a large share of the population suffered, and many of them died without proper medical supports.10 Improper synchronization among the responsible bodies, the Ministry of Health and Family Welfare (MoHFW), the Directorate General of Health Services, and test laboratories’ executives and workers, could be a reason for such mismanagements.11After detecting the first COVID-19 case on 8 March, in the following 2 weeks, more than 0.63 million overseas workers entered the country without proper screening, facilitating community transmission.6 But it is a matter of concern how effective these measures could be amid the intensifying nationwide corruption. Whereas mismanagements were paralyzing the health sector, increased corruption worsened the situation to a greater degree.6 The media reported 218 relief-related corruption incidents from March 10, 2020 to June 15, 2020, and most of the convicts were the government officials, public representatives, and ruling party leaders and activists.12 They either expropriated the public’s relief or counterfeited the budgets of medical equipment and health services.12 In July, a hospital owner along with a few responsible persons was convicted for trading with fake COVID-19 test certificates . More investigations reveal similar incidents, including the case of two doctors who threw thousands of patients’ samples in the garbage and provided them imaginary results  and kits, and their uneven distributions across the country, expensive coronavirus tests in private medicals (USD 50–60/test), the fewer number of medical workers, and unregulated testing system (elites get preferences).11 Moreover, many testing kits were preserved by corrupt businessmen to initiate an artificial crisis.16 Recently, the MoHFW imposed a fee on COVID-19 test in state-run laboratories too that dramatically reduced the average per day tests.17 Thanks to the lower test rate, 1,010 people died undiagnosed with coronavirus symptoms from March 2020 to May 2020.18 It hints about the possible discrepancies in the coronavirus’s official statistics of Bangladesh. Medical facilities, such as beds, intensive care units, and ventilators, are far fewer than the required amount in both government and private hospitals.19 Therefore, to manage a seat in the country’s finest hospitals, patients often need to have connections. Also, many patients prefer to remain at home fearing maltreatment in hospitals.10 A report reveals that 79% of patients stay at home and get treatment over the phone.20 In Bangladesh, only 3.05 physicians and 1.07 nurses serve every 10,000 people on average,5 which is insufficient for the pandemic situation. Moreover, medical workers were provided lower quality medical equipment, such as masks and personal protective equipment. Consequently, many doctors got infected and some died, making the doctors’ mortality rate of Bangladesh highest in the world.21Healthcare preparation and capacity against COVID-19 might explain the pandemic situation in Bangladesh more precisely. The coronavirus testing rate in Bangladesh (0.34%) is the second lowest in South Asia only after Afghanistan, a war-torn nation .6 Its mahttp://bdfactcheck.com, a nonprofit award-winning fact-checking website of Bangladesh). As a timely step, the government started detaining rumor-producers and rumor-spreaders to reduce the COVID-19 crisis to some degree. However, along with the perpetrators, as many human rights activists and organizations believe thanks to a few recent incidents, political dissidents and the government’s critics may be suppressed.23 Meanwhile, government officials are ordered “not to like, share or comment on social media posts” that criticize the government’s policies.22 These steps may breed fear among the online communities and hamper the positive health-related communication.Inadequate information flow and communication networks make the healthcare system more vulnerable and incompetent. Because of health-related uncertainty, information scarcity, the absence of reliable information sources, and defected flow of reliable information amid the pandemic, rumors became prevalent. In such a situation, national media outlets failed to successfully deliver reliable information to a large number of audiences, letting the more personalized and Internet-based media occupy the communication space. As a result, around 200 COVID-19–related online rumors spread across the country from March 2020 to July 2020 (for the complete list of rumors, see 24 This situation may be controlled by taking a few steps. First, corruption in the health sector is mandatory as this will help improve the proper utilization of allocated resources. Second, more tests should be conducted to identify the infected persons to provide them better treatment. Third, hospitals should be well equipped with updated and efficient medical supplies such as oxygen and medications to provide supportive treatment for COVID-19. Fourth, doctors and other medical workers must be protected from infection. Moreover, infected doctors and nurses could be super-spreaders of the virus. Fifth, thanks to higher population density and lower health awareness, social distancing in public spaces is virtually impossible in Bangladesh. Therefore, some sort of strict regulations for such spaces may be imposed. Sixth, healthy information flow is a must for the current COVID-19 situation in Bangladesh to reduce the health-related confusions and uncertainties. Battling the COVID-19 pandemic without organized strategies and an effective healthcare system would be like an attempt to kill a lion with bare hands.In June 2020 after a visit, a team of Chinese physicians expressed their concerns about Bangladesh’s disorganized health sector: this article already discussed the key selected discrepancies.