Knockdown of long non-coding RNA HOTAIR increases cisplatin sensitivity in ovarian cancer by inhibiting cisplatin-induced autophagy.

PURPOSE: Ovarian cancer is one of the most malignant tumors in the female reproductive system. With the widespread application of chemotherapeutic drugs, many ovarian cancer patients develop drug resistance. The aim of this study was to explore the function of HOTAIR in the treatment of ovarian cancer with cisplatin and its underlying mechanism. METHODS: Quantitative real-time polymerase chain reaction (qRT-PCR) was used to detect HOTAIR expressions in tissues and cells of ovarian cancer. Cell counting kit-8 (CCK-8) assay was used to examine the viability of ovarian cancer cells. Besides, small interfering (si) RNA was transfected to knockdown HOTAIR so as to explore its biological function. Western blot was performed to detect the expression levels of autophagy-related proteins. Flow cytometry was applied to detect apoptosis of ovarian cancer cells. RESULTS: HOTAIR was upregulated in ovarian cancer. Meanwhile, expression levels of autophagy-related proteins Atg7 and LC3 II/I in ovarian cancer cells increased with the increase of cisplatin concentration. Transfection of si-Atg7 could improve the therapeutic effect of cisplatin on ovarian cancer via inhibiting autophagy. Additionally, HOTAIR knockdown could increase the sensitivity of cisplatin in ovarian cancer treatment by inhibiting cisplatin-induced autophagy. CONCLUSIONS: Knockdown of long non-coding (lnc) RNA HOTAIR could increase the sensitivity of cisplatin in ovarian cancer by inhibiting cisplatin-induced autophagy. Our research attempts to find a more effective treatment and provides new ideas for the clinical treatment of ovarian cancer.

Ultrasound/CT combined with serum CEA/CA19.9 in the diagnosis and prognosis of rectal cancer.

PURPOSE: To explore the significance of computed tomography (CT) and transrectal ultrasonography (TRUS) combined with serum CEA and CA19.9 in the staging, diagnosis and prognosis of rectal cancer. METHODS: Fifty-six patients with rectal cancer were recruited from our oncology department. ELISA detected the expression level of CEA and CA19.9 in serum. The hemodynamic parameters of the rectal mucosa and tumor were detected by TRUS [resistance index (RI), pulse index (PI), peak systolic velocity (PSV), end-diastolic volume (EDV)]. All patients were pathologically examined to determine the disease stage and to compare the diagnostic accuracy of serum tumor markers, CT and TRUS. All patients were followed up for 24 months to assess the relationship between the combined examinations and the disease prognosis. RESULTS: CEA and CA19.9 levels were significantly different in patients with different pathological stages (p<0.05). RI and PI decreased with increasing pathological stage, while PSV and EDV were increased with increasing pathological stage. The serum CEA+CA19.9 examination showed 12 cases of misdiagnosis, with an accuracy diagnostic rate of 78.57% (447sol;56). CT examination showed 8 cases of misdiagnosis, with an accuracy diagnostic rate of 85.71% (48/56). TRUS showed 6 cases of misdiagnosis, with an accuracy diagnostic rate of 89.28% (50/56). However, only 2 cases were misdiagnosed and 96.43% (54/56) were accurate, while no statistical difference was noticed between combined detection and pathology (p<0.05). Postoperative follow-up showed significant differences in T staging at 6 months, 1 year and 2 years after operation (p<0.05). CONCLUSION: CT and TRUS combined with serum CEA and CA19.9 had great value in the diagnosis and prognosis in rectal cancer.

An investigation into the correlation between visual performance in simulated complex environments and academic attainment among primary school students.

Traditional vision screenings in schools are limited to simple visual tasks, yet students in their daily learning face more complex visual environments. Binocular rivalry tasks can partially simulate the visual challenges of real visual environments and activate advanced visual processing mechanisms that simple visual tasks cannot. Therefore, by superimposing binocular rivalry-state tasks onto simple visual tasks, we have developed an innovative vision screening program to rapidly and extensively assess students' visual performance in complex environments. This is a cross-sectional study in which we investigated the performance of 1126 grade 1-6 students from a primary school in Wuxi, China, in rivalry-state stereoscopic vision tasks. The correlation between the screening results of 1044 students and their academic achievements was also statistically analyzed. The study results revealed pass rates of 53.5-60.5% across various visual tests. Specifically, for first-grade students, there was a statistically significant difference in standardized Chinese scores between the group that failed and the group that passed the rivalry-state stereoscopic vision test (- 0.49 ± 3.42 vs. 0.22 ± 0.58, t = - 2.081, P = 0.04). This result underscores the importance of focusing on the visual adaptability of first graders in complex environments.Trail registration: Ethics Committee of Affiliated Children's Hospital of Jiangnan University-Certificate number: WXCH2022-04-027.

LCDM medium supports the derivation of bovine extended pluripotent stem cells with embryonic and extraembryonic potency in bovine-mouse chimeras from iPSCs and bovine fetal fibroblasts.

Cattle have emerged as one of the most important domestic animals widely used for meat, milk, and fur. Derivation of bovine pluripotent stem cells (PSCs) can be applied in drug selecting and human disease modeling and facilitated agriculture-related applications such as production of genetically excellent cattle by gene editing. Extended PSCs (EPSCs), capable of differentiating into embryonic and extraembryonic parts, have been generated in mouse, human, and pig. Whether bovine EPSCs could be generated, and their chimeric competency remains unclear. This study focused on derivation of bovine EPSCs using LCDM medium and exploring the characteristics of EPSCs among different species, including bovine, mouse, and human EPSCs. Here, using LCDM medium (consisting of hLIF, CHIR99021, (S)-(+)-dimethindene maleate, and minocycline hydrochloride) enables the derivation of bovine EPSCs from induced PSCs (iPSCs) and bovine fetal fibroblasts (BFF) with stable morphology, pluripotent marker expression, and in vitro differentiation ability. Notably, bovine EPSCs exhibited interspecies chimeric contribution to embryonic and extraembryonic tissues in pre-implantation blastocysts and postimplantation bovine-mouse chimeras. Transcriptome analysis revealed the unique molecular characteristics of bovine EPSCs compared with iPSCs. The similarities and differences in molecular features across bovine, human, and mouse EPSCs were also described by transcriptome analysis. Taken together, the LCDM culture system containing chemical cocktails can be used for the establishment and long-term passaging of bovine EPSCs with embryonic and extraembryonic potency in bovine-mouse chimeras. Our findings lay the foundation of generating PSCs in domestic animals and open avenues for basic and applied research in biology, medicine, and agriculture. DATABASE: Gene expression data of bovine EPSCs and bovine iPSCs are available in the GEO databases under the accession number PRJNA693452.

Neutrophil extracellular traps released by neutrophils impair revascularization and vascular remodeling after stroke.

Neovascularization and vascular remodeling are functionally important for brain repair after stroke. We show that neutrophils accumulate in the peri-infarct cortex during all stages of ischemic stroke. Neutrophils producing intravascular and intraparenchymal neutrophil extracellular traps (NETs) peak at 3-5 days. Neutrophil depletion reduces blood-brain barrier (BBB) breakdown and enhances neovascularization at 14 days. Peptidylarginine deiminase 4 (PAD4), an enzyme essential for NET formation, is upregulated in peri-ischemic brains. Overexpression of PAD4 induces an increase in NET formation that is accompanied by reduced neovascularization and increased BBB damage. Disruption of NETs by DNase 1 and inhibition of NET formation by genetic ablation or pharmacologic inhibition of PAD increases neovascularization and vascular repair and improves functional recovery. Furthermore, PAD inhibition reduces stroke-induced STING-mediated production of IFN-ß, and STING knockdown and IFN receptor-neutralizing antibody treatment reduces BBB breakdown and increases vascular plasticity. Collectively, our results indicate that NET release impairs vascular remodeling during stroke recovery.

[Epidemiologic study of dry eye in populations equal or over 20 years old in Jiangning District of Shanghai].

OBJECTIVE: To investigate the prevalence of dry eye in populations equal or over 20 years old in Jiangning District, Shanghai, China. METHODS: This was a cross-sectional study. From September 2008 to January 2009, 6 small districts including 21,102 people of Jiangning District were randomly selected as survey venues by Department of Ophthalmology in First People's Hospital affiliated to Shanghai Jiaotong University. Then, 1266 people as the selected residents were enrolled, which was figured out through the random cluster sampling procedure. Every participant completed dry eye questionnaire, the ocular surface disease index (OSDI), and a series of examination including slit-lamp microscope, tear-film break-up time (BUT) , Schirmer I test, and fluorescein staining of the cornea (F1). The diagnosis of dry eye was referred to the well-accepted domestic diagnostic criteria The SPSS11. 0 software was used to analyze the database, t . test, chi2 test, one-way-ANOVA and Logistic regression were used for analysis. RESULTS: One thousand and eighty five residents finally took part in this study, and the inclusion ratio was 85.70%. Three hundred and twenty six individuals, including 101 men and 225 women, were diagnosed as dry eye, and the prevalence rate was 30.05%. The prevalence of dry eye in the female (33.78%) was higher than that of the male (24.11%) (chi2 = 11.46, P < 0.01). The prevalence of dry eye in people over 50 years old was higher than that under 50 years (chi2 = 94.50, P < 0.01). The figure of Schirmer I test and BUT decreased in elder people, at the same time the scores of Fl and MGD increased. Meanwhile, the score of OSDI in dry eye patients was significantly higher than that in non-dry eye individuals. The relative risk factors of dry eye were gender, age, wearing contact lens, long-time using of eye solutions, taking anti-allergy drugs. CONCLUSIONS: The prevalence of dry eye in female is higher than that in the male. And the prevalence of dry eye increases following the aging process. Relative risk factors of dry eye are gender, age, wearing contact lens, long-time using of eye solutions, taking anti-allergy drugs.

[Applications of mass spectrometry-based proteomics in food authentication and quality identification].

Proteomics, as a new research direction in the post-genomics era, has been developing rapidly in recent years and has been applied in many fields, becoming a powerful research tool in food quality identification and safety control. Proteomics opens up a new horizon in food science research. It can not only identify the species of characteristic proteins, but also quantify the response of targeted proteins. Proteomics allows for the dynamic analysis of protein composition and content in samples that vary in species, geographical origin or growth stage. The research methods of proteomics are varied, and mass spectrometry (MS) is one of the most common techniques. This paper introduces the concept, classification, and research technology of proteomics, as well as common protein databases. Applications of MS-based proteomics in food authentication and quality identification were reviewed, including seafood, meat products, dairy products, health food, and value-added food. Furthermore, the future development of proteomics was investigated.

Evolution of Grain Interfaces in Annealed Duplex Stainless Steel after Parallel Cross Rolling and Direct Rolling.

Changes in various grain interfaces, including the grain boundary and phase boundary, are a strong indication of microstructural changes, particularly ultra-fined grains achieved by large strain deformation and subsequent annealing. After direct rolling and cross rolling with the same strain of ε = 2, the distributions of the interfaces in annealed UNS S32304 duplex stainless steel were investigated using electron backscatter diffraction (EBSD) in this study. The ferrite experienced continued recovery, and a high density of low-angle grain boundaries (LAGBs) was produced. The percentage and number of twin boundaries (TBs) and LAGBs varied within the austenite. TBs were frequently found within austenite, showing a deviation from the Kurdjumov-Sachs (K-S) orientation relationship (OR) with ferrite matrix. However, LAGBs usually occur in austenite, with the K-S OR in the ferrite matrix. LAGBs were prevalent in the precipitated austenite grains, and therefore a strong texture was introduced in the cross-rolled and annealed samples, in which the precipitated austenite readily maintained the K-S OR in the ferrite matrix. By contrast, more TBs and a less robust texture were found in the precipitated austenite in direct-rolled and annealed samples, deviating from the K-S OR.

Activation of Transient Receptor Potential Vanilloid 4 Promotes the Proliferation of Stem Cells in the Adult Hippocampal Dentate Gyrus.

Neurogenesis plays an important role in adult hippocampal function, and this process can be modulated by intracellular calcium. The activation of transient receptor potential vanilloid 4 (TRPV4) induces an increase in intracellular calcium concentration, but whether neurogenesis can be modulated by TRPV4 activation remains unclear. Here, we report that intracerebroventricular injection of the TRPV4 agonist GSK1016790A for 5 days enhanced the proliferation of stem cells in the hippocampal dentate gyrus (DG) of adult mice without affecting neurite growth, differentiation, or survival of newborn cells. GSK1016790A induced increases in the hippocampal protein levels of cyclin-dependent kinase (CDK) 6, CDK2, cyclin E1, and cyclin A2 but did not affect CDK4 and cyclin D1 expression. The phosphorylation of retinoblastoma protein (Rb) in hippocampi was enhanced in GSK1016790A-injected mice compared with control mice. Moreover, hippocampal protein levels of extracellular signal-regulated kinase 1/2 (ERK1/2) and p38 mitogen-activated protein kinase (p38 MAPK) phosphorylation were enhanced by GSK1016790A. Finally, GSK1016790A-enhanced proliferation was markedly blocked by a MAPK/ERK kinase or p38 MAPK antagonist (U0126 or SB203580, respectively). The increased protein levels of CDK2 and CDK6, as well as those of cyclin E1 and cyclin A2, in GSK1016790A-injected mice were substantially reduced by co-injection of U0126 or SB203580. We conclude that TRPV4 activation results in the proliferation of stem cells in the adult hippocampal DG, which is likely mediated through ERK1/2 and p38 MAPK signaling to increase the expression of CDKs (CDK6 and CDK2) and cyclins (cyclin E1 and A2), phosphorylate Rb consequently, and accelerate the cell cycle ultimately.

Activation of Transient Receptor Potential Vanilloid 4 Impairs the Dendritic Arborization of Newborn Neurons in the Hippocampal Dentate Gyrus through the AMPK and Akt Signaling Pathways.

Neurite growth is an important process for the adult hippocampal neurogenesis which is regulated by a specific range of the intracellular free Ca2+ concentration ([Ca2+]i). Transient receptor potential vanilloid 4 (TRPV4) is a calcium-permeable channel and activation of it causes an increase in [Ca2+]i. We recently reported that TRPV4 activation promotes the proliferation of stem cells in the adult hippocampal dentate gyrus (DG). The present study aimed to examine the effect of TRPV4 activation on the dendrite morphology of newborn neurons in the adult hippocampal DG. Here, we report that intracerebroventricular injection of the TRPV4 agonist GSK1016790A for 5 days (GSK1016790A-injected mice) reduced the number of doublecortin immunopositive (DCX+) cells and DCX+ fibers in the hippocampal DG, showing the impaired dendritic arborization of newborn neurons. The phosphorylated AMP-activated protein kinase (p-AMPK) protein level increased from 30 min to 2 h, and then decreased from 1 to 5 days after GSK1016790A injection. The phosphorylated protein kinase B (p-Akt) protein level decreased from 30 min to 5 days after GSK1016790A injection; this decrease was markedly attenuated by the AMPK antagonist compound C (CC), but not by the AMPK agonist AICAR. Moreover, the phosphorylated mammalian target of rapamycin (mTOR) and p70 ribosomal S6 kinase (p70S6k) protein levels were decreased by GSK1016790A; these changes were sensitive to 740 Y-P and CC. The phosphorylation of glycogen synthase kinase 3ß (GSK3ß) at Y216 was increased by GSK1016790A, and this change was accompanied by increased phosphorylation of microtubule-associated protein 2 (MAP2) and collapsin response mediator protein-2 (CRMP-2). These changes were markedly blocked by 740 Y-P and CC. Finally, GSK1016790A-induced decrease of DCX+ cells and DCX+ fibers was markedly attenuated by 740 Y-P and CC, but was unaffected by AICAR. We conclude that TRPV4 activation impairs the dendritic arborization of newborn neurons through increasing AMPK and inhibiting Akt to inhibit the mTOR-p70S6k pathway, activate GSK3ß and thereby result in the inhibition of MAP2 and CRMP-2 function.

Transient Receptor Potential Vanilloid 4-Induced Modulation of Voltage-Gated Sodium Channels in Hippocampal Neurons.

Transient receptor potential vanilloid 4 (TRPV4) is reported to control the resting membrane potential and increase excitability in many types of cells. Voltage-gated sodium channels (VGSCs) play an important role in initiating action potentials in neurons. However, whether VGSCs can be modulated by the activation of TRPV4 in hippocampal pyramidal neurons remains unknown. In this study, we tested the effect of TRPV4 agonists (GSK1016790A and 4α-PDD) on voltage-gated sodium current (I Na) in hippocampal CA1 pyramidal neurons and the protein levels of α/ß-subunit of VGSCs in the hippocampus of mice subjected to intracerebroventricular (icv.) injection of GSK1016790A (GSK-injected mice). Herein, we report that I Na was inhibited by acute application of GSK1016790A or 4α-PDD. In the presence of TRPV4 agonists, the voltage-dependent inactivation curve shifted to the hyperpolarization, whereas the voltage-dependent activation curve remained unchanged. The TRPV4 agonist-induced inhibition of I Na was blocked by the TRPV4 antagonist or tetrodotoxin. Moreover, blocking protein kinase A (PKA) markedly attenuated the GSK1016790A-induced inhibition of I Na, whereas antagonism of protein kinase C or p38 mitogen-activated protein kinase did not change GSK1016790A action. Finally, the protein levels of Nav1.1, Nav1.2, and Nav1.6 in the hippocampus increased in GSK-injected mice, whereas those of Nav1.3 and Navß1 remained nearly unchanged. We conclude that I Na is inhibited by the acute activation of TRPV4 through PKA signaling pathway in hippocampal pyramidal neurons, but protein expression of α-subunit of VGSCs is increased by sustained TRPV4 activation, which may compensate for the acute inhibition of I Na and provide a possibility for hyper-excitability upon sustained TRPV4 activation.

Enhanced Oxidative Stress Is Responsible for TRPV4-Induced Neurotoxicity.

Transient receptor potential vanilloid 4 (TRPV4) has been reported to be responsible for neuronal injury in pathological conditions. Excessive oxidative stress can lead to neuronal damage, and activation of TRPV4 increases the production of reactive oxygen species (ROS) and nitric oxide (NO) in many types of cells. The present study explored whether TRPV4-induced neuronal injury is mediated through enhancing oxidative stress. We found that intracerebroventricular injection of the TRPV4 agonist GSK1016790A increased the content of methane dicarboxylic aldehyde (MDA) and NO in the hippocampus, which was blocked by administration of the TRPV4 specific antagonist HC-067047. The activities of catalase (CAT) and glutathione peroxidase (GSH-Px) were decreased by GSK1016790A, whereas the activity of superoxide dismutase (SOD) remained unchanged. Moreover, the protein level and activity of neuronal nitric oxide synthase (nNOS) were increased by GSK1016790A, and the GSK1016790A-induced increase in NO content was blocked by an nNOS specific antagonist ARL-17477. The GSK1016790A-induced modulations of CAT, GSH-Px and nNOS activities and the protein level of nNOS were significantly inhibited by HC-067047. Finally, GSK1016790A-induced neuronal death and apoptosis in the hippocampal CA1 area were markedly attenuated by administration of a ROS scavenger Trolox or ARL-17477. We conclude that activation of TRPV4 enhances oxidative stress by inhibiting CAT and GSH-Px and increasing nNOS, which is responsible, at least in part, for TRPV4-induced neurotoxicity.

Transient Receptor Potential Vanilloid 4 Inhibits γ-Aminobutyric Acid-Activated Current in Hippocampal Pyramidal Neurons.

The balance between excitatory and inhibitory neurotransmitter systems is crucial for the modulation of neuronal excitability in the central nervous system (CNS). The activation of transient receptor potential vanilloid 4 (TRPV4) is reported to enhance the response of hippocampal glutamate receptors, but whether the inhibitory neurotransmitter system can be regulated by TRPV4 remains unknown. γ-aminobutyric acid (GABA) is the major inhibitory neurotransmitter in the CNS. Here, we show that application of transient receptor potential vanilloid 4 (TRPV4) synthetic (GSK1016790A or 4α-PDD) or endogenous agonist (5,6-EET) inhibited GABA-activated current (I GABA) in hippocampal CA1 pyramidal neurons, which was blocked by specific antagonists of TRPV4 and of GABAA receptors. GSK1016790A increased the phosphorylated AMP-activated protein kinase (p-AMPK) and decreased the phosphorylated protein kinase B (p-Akt) protein levels, which was attenuated by removing extracellular calcium or by a calcium/calmodulin-dependent protein kinase kinase-ß antagonist. GSK1016790A-induced decrease of p-Akt protein level was sensitive to an AMPK antagonist. GSK1016790A-inhibited I GABA was blocked by an AMPK antagonist or a phosphatidyl inositol 3 kinase (PI3K) agonist. GSK1016790A-induced inhibition of I GABA was also significantly attenuated by a protein kinase C (PKC) antagonist but was unaffected by protein kinase A or calcium/calmodulin-dependent protein kinase II antagonist. We conclude that activation of TRPV4 inhibits GABAA receptor, which may be mediated by activation of AMPK and subsequent down-regulation of PI3K/Akt signaling and activation of PKC signaling. Inhibition of GABAA receptors may account for the neuronal hyperexcitability caused by TRPV4 activation.

Blockage of transient receptor potential vanilloid 4 inhibits brain edema in middle cerebral artery occlusion mice.

Brain edema is an important pathological process during stroke. Activation of transient receptor potential vanilloid 4 (TRPV4) causes an up-regulation of matrix metalloproteinases (MMPs) in lung tissue. MMP can digest the endothelial basal lamina to destroy blood brain barrier, leading to vasogenic brain edema. Herein, we tested whether TRPV4-blockage could inhibit brain edema through inhibiting MMPs in middle cerebral artery occlusion (MCAO) mice. We found that the brain water content and Evans blue extravasation at 48 h post-MCAO were reduced by a TRPV4 antagonist HC-067047. The increased MMP-2/9 protein expression in hippocampi of MCAO mice was attenuated by HC-067046, but only the increased MMP-9 activity was blocked by HC-067047. The loss of zonula occludens-1 (ZO-1) and occludin protein in MCAO mice was also attenuated by HC-067047. Moreover, MMP-2/9 protein expression increased in mice treated with a TRPV4 agonist GSK1016790A, but only MMP-9 activity was increased by GSK1016790A. Finally, ZO-1 and occludin protein expression was decreased by GSK1016790A, which was reversed by an MMP-9 inhibitor. We conclude that blockage of TRPV4 may inhibit brain edema in cerebral ischemia through inhibiting MMP-9 activation and the loss of tight junction protein.

Cooperative complexation of amino acid derivatives to platinum acetylide-based bolaamphiphile.

A platinum(II) acetylide-based bolaamphiphile equipped with two peripheral B18C6 moieties has been successfully prepared, which demonstrates cooperative recognition behavior toward l-alanine ester salt in chloroform. In a polar methanol/chloroform (3/1, v/v) medium, the amino acid additives influence the aggregation of the bolaamphiphile significantly, leading to the morphological transition from nanospherical to disordered structures. The adaptive properties of the current host-guest binary system will benefit the development of stimuli-responsive supramolecular materials.

Cooperative self-assembly of platinum(II) acetylide complexes.

Rod-like platinum(II) acetylide complexes have been demonstrated to form one-dimensional helical supramolecular polymers by the cooperative growth mechanism, leading to supramolecular gels by bundling single fibrils into entangled networks.

Formation of stimuli-responsive supramolecular polymeric assemblies via orthogonal metal-ligand and host-guest interactions.

Linear supramolecular polymers, assembled via the combination of orthogonal terpyridine-Zn(2+) and benzo-21-crown-7/secondary ammonium salt recognition motifs, exhibit dynamic properties responsive to various external stimuli.

Comparative characterization of chitinases from silkworm (Bombyx mori) and bollworm (Helicoverpa armigera).

Chitinases play an important role in the degradation of the cuticular chitin during the process of ecdysis. In this study, we compared the chitinases of two insect species, Bombyx mori (silkworm) and Helicoverpa armigera (bollworm), to assess the relation between characteristics and chitinase patterns. Differences between two chitinases were observed after purification using ammonium sulfate precipitation, affinity chromatography, and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) assay. Although the specific activities of the purified enzymes were different, the purification yields were similar. One band of 88 kDa was observed for B. mori, and the other band of 75 kDa was detected for H. armigera. When a range of properties was tested, it was found that the optimum temperatures of B. mori and H. armigera chitinases were 45 and 50°C, respectively; the optimum pH value was 6.0 for both chitinases. Mn(2+) played catalytic role while Cu(2+) and SDS strongly inhibited activities of both enzymes. Between two chitinases, differences in K (M) were also observed. K (M) of chitinase from silkworm and bollworm was found to be 22.3 and 41.0 µmol/l, respectively. Both the chitinases significantly inhibited the spore germination of two fungal species, Saccharomyces cerevisiae and Penicillium.