Manipulation of isocitrate dehydrogenase genes affects the anti-autolytic ability of lager yeast / 生物工程学报

Yeast autolysis affects the flavor and quality of beer. The regulation of yeast autolysis is a need for industrial beer production. Previous studies on brewer's yeast autolysis showed that the citric acid cycle-related genes had a great influence on yeast autolysis. To explore the contribution of isocitrate dehydrogenase genes in autolysis, the IDP1 and IDP2 genes were destroyed or overexpressed in typical lager yeast Pilsner. The destruction of IDP1 gene improved the anti-autolytic ability of yeast, and the anti-autolytic index after 96 h autolysis was 8.40, 1.5 times higher than that of the original strain. The destruction of IDP1 gene increased the supply of nicotinamide adenine dinucleotide phosphate (NADPH) and the NADPH/NADP+ ratio was 1.94. After fermentation, intracellular ATP level was 1.8 times higher than that of the original strain, while reactive oxygen species (ROS) was reduced by 10%. The destruction of IDP2 gene resulted in rapid autolysis and a decrease in the supply of NADPH. Anti-autolytic index after 96 h autolysis was 4.03 and the NADPH/NADP+ ratio was 0.89. After fermentation, intracellular ATP level was reduced by 8% compared with original strain, ROS was 1.3 times higher than that of the original strain. The results may help understand the regulation mechanism of citric acid cycle-related genes on yeast autolysis and provide a basis for the selection of excellent yeast with controllable anti-autolytic performance.

Regulations of RLM1 gene affect the anti-autolytic ability of lager yeast / 生物工程学报

The autolysis of brewer's yeast seriously affects the quality of beer and the quality of yeast is considered as one of the key factors in beer brewing. Previous studies on brewer's yeast autolysis showed that RLM1 gene, an important transcription factor in cell integrity pathway, is closely related to the autolysis of yeast. In this study, RLM1 was knocked out and overexpressed in a haploid brewer's yeast. RLM1 disruption resulted in poor anti-autolysis performance of yeast, whereas overexpression of RLM1 contributed to the anti-autolytic ability of yeast. In addition, RLM1 gene knockout affected the osmotic stress resistance, cell wall damage resistance, nitrogen starvation resistance and temperature tolerance of yeast strain. The transcriptional level of GAS1 involved in cell wall assembly and DNA damage response was regulated along with the expression of RLM1, whereas other genes in CWI pathway did not show apparent regularity. RLM1 might mainly affect the expression of GAS1 so as to improve the stress resistance of lager yeast in harsh environment. The result from this study help further understand the mechanism of yeast autolysis and lay a foundation for breeding brewer's yeast strain with better anti-autolytic ability.

Optimization of yeast autolysis under solid-state fermentation conditions / 生物工程学报

Yeast autolysis under solid-state fermentation can effectively promote the release of various active substances, thereby improving the quality of yeast products. The optimal process for yeast autolysis under solid-state fermentation was obtained by optimizing the autolysis temperature, autolysis time and the zinc ion concentration. We analyzed the indexes of free amino acid, soluble protein and α-amino nitrogen in the fermentation material, as well as A₂₆₀/A₂₈₀ ratio to determine yeast autolysis process conditions in the solid-state fermentation. On the basis of the obtained data, L₉ (3³) orthogonal test was designed to optimize the solid-state fermentation parameters for yeast autolysis: temperature at 40, 50 and 55 °C; time 12, 18 and 24 h; zinc ion concentration 2, 4 and 8 mg/kg. The optimum process conditions for yeast autolysis were: autolysis temperature 55 °C, time 18 h, zinc ion concentration 2 mg/kg, and soluble protein content reached 9.31 mg/g, free amino acid 14.36 mg/g, α-amino nitrogen 10.16 μg/g and A₂₆₀/A₂₈₀ 1.73. After optimization of the process, the soluble protein, free amino acid and α-amino nitrogen contents of the yeast autolysis production can be significantly increased, thereby obviously improving the quality of the composite culture.

Usefulness of Dipstick Test for Vitreous Glucose in Autopsy Practice / 대한법의학회지

The concentration of glucose in the vitreous humor serves as an important diagnostic marker for diabetic mellitus in post-mortem examinations, as the vitreous humor can be easily collected and the glucose test using vitreous humor is not significantly affected by cell autolysis and hemolysis. For a quick and effective glucose test, we suggest a dipstick test of the vitreous humor during autopsy. The results were evaluated and compared with other methods for significance testing. In this study, vitreous humor was analyzed from 257 autopsy cases. Qualitative concordance rate of the dipstick test for glucose and the hexokinase test was 98.7%, positive prediction rate was 89.6%, and negative prediction rate was 100%. However, there was no significant correlation between the dipstick glucose test and the hexokinase test. We conclude that the dipstick glucose test is effective and useful for post-mortem glucose screening testing and for additional post-mortem diabetes testing. Recently, the importance of post-mortem glucose testing has increased with the increase in deaths from diabetes complications. The use of the dipstick glucose test in autopsy practice can improve forensic medicine in Korea.

Acinar autolysis and mucous extravasation in human sublingual glands: a microscopic postmortem study

Although some morphological investigations on aged human sublingual glands (HSG) found eventual phenomena identified as autolysis and mucous extravasation, the exact meaning of these findings has not been elucidated.Objective The aim of this work is to investigate whether acinar autolysis and mucous extravasation are related to the aging process in human sublingual glands. We also speculate if autolytic changes may assist forensic pathologists in determining time of death.Material and Methods 186 cadavers’ glands were allocated to age groups: I (0–30 years); II (31–60), and III (61–90). Time and mode of death were also recorded. Acinar autolysis and mucous extravasation were classified as present or absent. Ultrastructural analysis was performed using transmission electron microscopy (TEM). Data were compared using Mann-Whitney U, Spearman’s correlation coefficient, Kruskal-Wallis, and Dunn tests (p<0.05).Results There was correlation between age and acinar autolysis (r=0.38; p=0.0001). However, there was no correlation between autolysis and time of death. No differences were observed between genders. TEM showed mucous and serous cells presenting nuclear and membrane alterations and mucous cells were more susceptible to autolysis.Conclusion Acinar autolysis occurred in all age groups and increased with age while mucous extravasation was rarely found. Both findings are independent. Autolysis degrees in HSG could not be used to determine time of death.

Overexpression of FKS1 to improve yeast autolysis-stress / 生物工程学报

With the development of high gravity brewing, yeast cells are exposed to multiple brewing-associated stresses, such as increased osmotic pressure, enhanced alcohol concentration and nutritional imbalance. These will speed up yeast autolysis, which seriously influence beer flavor and quality. To increase yeast anti-autolytic ability, FKS1 overexpression strain was constructed by 18S rDNA. The concentration of β-1,3-glucan of overexpression strain was 62% higher than that of wild type strain. Meantime, FKS1 overexpression strain increased anti-stress ability at 8% ethanol, 0.4 mol/L NaCl and starvation stress. Under simulated autolysis, FKS1 showed good anti-autolytic ability by slower autolysis. These results confirms the potential of FKS1 overexpression to tackle yeast autolysis in high-gravity brewing.

Histopathological analysis of organs submitted by legal medicine experts in Baojii City: 358 forensic identification cases / 法医学杂志

OBJECTIVE@#To analyze pathological characteristics of organs recovered during forensic autopsy submitted by legal medicine experts.@*METHODS@#From Baoji city, 358 cases of forensic autopsy specimens from a series of routine exams were collected. And histopathological diagnoses were reviewed.@*RESULTS@#Majority of the 358 cases were young men. The major causes of death were trauma, sudden death and poisoning. The cause of death was determined with histology in 250 cases. No typical histological changes were noted in 101 cases. The tissue autolysis and decomposition were present in 7 cases. The major pathological diagnosis was cardiovascular disease, followed by diseases in respiratory, nervous, and digestive systems.@*CONCLUSION@#Forensic autopsy with its professional characteristics, is different from regular autopsy. When diagnosing cause of death by histopathological examination, pathologists should collaborate with legal medicine experts to know the details of the cases, circumstances surrounding the death, and specific forensic pathological characteristics.

Discovery of a novel gene involved in autolysis of Clostridium cells

Cell autolysis plays important physiological roles in the life cycle of clostridial cells. Understanding the genetic basis of the autolysis phenomenon of pathogenic Clostridium or solvent producing Clostridium cells might provide new insights into this important species. Genes that might be involved in autolysis of Clostridium acetobutylicum, a model clostridial species, were investigated in this study. Twelve putative autolysin genes were predicted in C. acetobutylicum DSM 1731 genome through bioinformatics analysis. Of these 12 genes, gene SMB_G3117 was selected for testing the in tracellular autolysin activity, growth profile, viable cell numbers, and cellular morphology. We found that overexpression of SMB_G3117 gene led to earlier ceased growth, significantly increased number of dead cells, and clear electrolucent cavities, while disruption of SMB_G3117 gene exhibited remarkably reduced intracellular autolysin activity. These results indicate that SMB_G3117 is a novel gene involved in cellular autolysis of C. acetobutylicum.

Expression and Activity of Catalases Is Differentially Affected by GpaA (Ga) and FlbA (Regulator of G Protein Signaling) in Aspergillus fumigatus

Vegetative growth signaling of the opportunistic human pathogenic fungus Aspergillus fumigatus is mediated by GpaA (Galpha). FlbA is a regulator of G protein signaling, which attenuates GpaA-mediated growth signaling in this fungus. The flbA deletion (DeltaflbA) and the constitutively active GpaA (GpaA(Q204L)) mutants exhibit enhanced proliferation, precocious autolysis, and reduced asexual sporulation. In this study, we demonstrate that both mutants also show enhanced tolerance against H2O2 and their radial growth was approximately 1.6 fold higher than that of wild type (WT) in medium with 10 mM H2O2. We performed quantitative PCR (qRT-PCR) for examination of mRNA levels of three catalase encoding genes (catA, cat1, and cat2) in WT and the two mutants. According to the results, while levels of spore-specific catA mRNA were comparable among the three strains, cat1 and cat2 mRNA levels were significantly higher in the two mutants than in WT. In particular, the DeltaflbA mutant showed significantly enhanced and prolonged expression of cat1 and precocious expression of cat2. In accordance with this result, activity of the Cat1 protein in the DeltaflbA mutant was higher than that of gpaA(Q204L) and WT strains. For activity of the Cat2 protein, both mutants began to show enhanced activity at 48 and 72 hr of growth compared to WT. These results lead to the conclusion that GpaA activates expression and activity of cat1 and cat2, whereas FlbA plays an antagonistic role in control of catalases, leading to balanced responses to neutralizing the toxicity of reactive oxygen species.

Histologic Estimation of Intrauterine Retention Time after Fetal Death / 대한법의학회지

The intrauterine retention time (IURT) after fetal death can be estimated from the loss of nuclear basophilia. We therefore attempted to derive an autolysis equation to estimate IURT in experimental rat fetuses and human fetal autopsy slides. The degree of loss of nuclear basophilia in various tissues was assessed by hematoxylin and eosin (H & E) staining. Fetal rat tissues showed different rates of autolysis, allowing for the construction of an experimental autolysis curve. We also reviewed the H & E stained slides obtained from 27 human fetal autopsy cases with well-documented death intervals. The degree of autolysis in various tissues was evaluated using percentile scores (PS). Using the findings from H&E staining, we derived the equation Ln (PS/[100-PS]) = 2.62716-0.02377 x IURT. However, this equation or autolysis scores showed some limitations. Owing to the inconsistency of PS, this equation is reliably applicable only within 24 hours of intrauterine fetal death. In the fetal autopsy review, fetal hydrops, local effusion, and sepsis also contributed to accelerated autolysis.

Effect of pH and salt gradient on the autolysis of Lactococcus lactis strains

The aim of this work was to assess in-vitro the effect of pH and salt concentration on the rate of autolysis in L. lactis strains. Regardless autolysis variation among L. lactis strains, statistical analysis showed evidence of increase of autolysis in L. lactis under low salt concentration and acidic conditions.

Cristalino fagocitado: presentación de un caso / Phagocytic crystalline: a case presentation

El traumatismo ocular constituye uno de los mayores retos en la cirugía oftalmológica y a su vez es un gran simulador. Son múltiples las formas de presentación de una catarata traumática y la evolución de ésta en los días siguientes al trauma. En ocasiones a pesar de ser un trauma de gran magnitud, el cristalino y sus cápsulas quedan intactas, con el paso de los días comienza a opacificarse, produciéndose la catarata y la disminución de la agudeza visual. Caso clínico: se reporta el caso de un paciente masculino, blanco, de 56 años de edad, trabajador agrícola, con antecedentes de salud aparente, que recibió un trauma contuso en el ojo derecho a consecuencia de lo cual se produjo una catarata asociada a un proceso inflamatorio crónico, hecho que provocó la fagocitosis de una porción del cristalino, con la formación de sinequias posteriores además de neovascularización del iris y cristalino en la zona de las mismas. Se intervino quirúrgicamente. La evolución postoperatoria fue buena, no presentó complicaciones. La recuperación visual fue excelente. Conclusiones: la recuperación final en pacientes con catarata traumática y cristalino fagocitado puede ser satisfactoria.

The ocular traumatism constitutes one of the biggest challenges in ophthalmologic surgery and at the same time is a great simulator. There are multiple ways of presentation of a traumatic cataract and its evolution in the following days to a trauma. In occasions in spite of being a trauma of great magnitude, the lens of the eye and its capsulae are intact, as days goes by it begins an opacification, causing the cataract and reducing the visual acuity. Clinical case: a masculine, white, 56 years old patient, agricultural worker is presented, with apparent health antecedents which received a bruised trauma in the right eye and as a consequence a cataract associate to a chronic inflammatory process was originated, which caused phagocytosis of a portion of the crystalline, with the formation of posterior synechiae as well as neovascularization of the iris and crystalline. He was surgically intervened. No complications in the postoperative evolution. The visual recovery was excellent.

Postmortem acinar autolysis in rat sublingual gland: a morphometric study

ABSTRACT OBJECTIVE: To analyze and to quantify morphological acinar postmortem changes in rat sublingual glands (SLG). MATERIAL AND METHODSs: Fifty rats were divided into two groups of 25 animals each. Group I was used for morphological and morphometric evaluations and group II for the determination of gland density and processed gland volume. Acinar autolytic changes were studied at 0 (control group), 3, 6, 12 and 24 h postmortem periods. The morphometric analysis of the volume density (Vv) and total volume (Vt) of intact (ia) and autolyzed (aa) acini was performed under light microscopy using a Zeiss II integration grid with 100 symmetrically distributed points. RESULTS: Morphologically, temporal progressive nuclear alterations and gradual loss of the structural architecture of acinar cells were found. Regarding quantitative results, both the Vvaa and the Vvia showed statistically significant differences among all postmortem periods (p<0.05). Vvaa increased from 0.42 percent at 0 h to 75.84 percent at 24 h postmortem and Vvia decreased from 71.16 percent to 0 percent over the same period. For Vtaa and Vtia, no statistically significant differences occurred between 12-24 h and 0-3 h (p>0.05), respectively. Vtaa increased from 0.18 mm³ at 0 h to 38.17 mm³ at 12 h, while Vtia showed a decrease from 33.47 mm³ to 0 mm³ between 3-24 h postmortem. Data concerning Vtaa were adjusted by two-variable linear regression, obtaining the equation: y=-3.54 + 3.38x (r²=0.90). The Vtaa growth rate calculated by this equation was 3.38 mm³/h between 0-12 h. CONCLUSION: Acinar autolysis on rat SLG demonstrated the most significant signs during the first 6 h postmortem and was widely spread through the gland at 12 h.

Autólise da camada de células granulares do cerebelo em bovinos / Conglutination of the cerebellar granule cell layer in cattle

A autólise da camada de células granulares do cerebelo é um achado postmortem observado em bovinos e relacionado ao prolongado tempo de duração entre a morte e a fixação do encéfalo. Para observar a prevalência desse artefato, 228 cerebelos foram examinados histologicamente ao longo do verme cerebelar. Trinta e seis por cento desses casos apresentaram conglutinação cerebelar não relacionada a doença específica. Em 74,4 por cento dos casos em que foi observada conglutinação cerebelar, não foram observadas alterações histopatológicas. Histologicamente, a conglutinação cerebelar caracterizou-se por dissolução segmentar ou difusa da camada de células granulares. A descrição deste artefato é importante, pois já foi confundido como lesão relacionada a doença específica do sistema nervoso de bovinos.

Cerebellar conglutination of the granule cell layer is a postmortem artifact observed in cattle. It is related to increased time between death and fixation of the brain. In this study, 228 cerebellum were histologically analyzed throughout cerebellar vermis. Cerebellar conglutination was observed in 36 percent of the cases and it was not related to specific diseases. Seventy-four percent of the cases did not present any histological lesions in the brain. Microscopically, cerebellar conglutination was characterized by segmentar or diffuse dissolution of the granule cell layer. This artifact has been misinterpreted as resulting from specific diseases of the central nervous system in cattle, and it has to be differentiated from important antemortem lesions.

Protein content in irrigating solutions in contact with pulp tissue

Las soluciones de irrigación endodóntica pueden tener distintas accionesLas soluciones de irrigación endodóntica pueden tener distintas acciones. Una de ellas es la de disolver el tejido pulpar. Seevaluó in vitro la capacidad de diferentes soluciones de irrigaciónpara disolver tejido pulpar vital y necrótico mediante un estudio cuanti y cualitativo de proteínas pulpares totales solubles. Se utilizaron pulpas de dientes de bovinos jóvenes, vitales y con necrosis inducida. Se seccionaron en trozos pequeños, que se pesaron y colocaron en 1 ml de las siguientes soluciones: hipoclorito de sodio al 1por ciento y 2,5 por ciento, hidróxido de calcio al 1 por ciento y 5 por ciento, gluconato de clorhexidina al 0,2 por ciento, té al 1 por ciento y aguadestilada (control). Se colocaron a 37°C y se extrajeron muestras de 20 μl a luego de 30 min, 90 min y 20 hs. Se dosaron proteínas totales utilizando el método de Lowry y por electroforesis(SDS-Page 12 por ciento). Se determinaron bandas de proteínassolubles. Los resultados se analizaron por Anova En el analisis químico, de las corridas electroforéticas de proteínaspulpares bovinas se evidenció que tanto el hipoclorito de sodio en ambas concentraciones como el hidróxido de calcio producen desnaturalización de proteínas. No se demuestra acción solvente con clorhexidina, té y agua destilada.

Production of fish protein hydrolyzate for use as fortifying agent in some food products

Silver carp fish was used as a raw material for preparing protein hydrolyzate by two methods: 1] Autlysis in the presence of sodium chloride or without it at 65 °C and 2] Enzymatic hydrolysis using two proteolytic enzymes, pepsin or papain to obtain fish protein hydrolyzate under controlled conditions. The obtained papain protein hydrolyzate had high quantity and quality as compared with the other produced hydrolyzates. The obtained hydrolyzate was used for fortification of Egyptian shamy bread and biscuit as source of essential amino acids. It was added in different levels [2-4%] for shamy bread and [0.5-5.0%] for biscuits. Chemical, nutritional and organoleptic properties of supplemented shamy bread and biscuit were evaluated and the results were as follows: 1-Produced silver carp protein hydrolyzates contained [64.69-93.9 1%] protein, [0.44-0.87%] fat, [4.91-33.01%] ash and [0.54-3.37%] carbohydrates [on dry weight basis] depending on the method of hydrolysis used. Also, the amino acid composition of papain fish protein hydrolyzate contained the following amino acids [gm/100 gm protein]: Isoleucine 4.72, leucine 8.64, lysin 8.74, threonine 4.67, valine 4.96, histidin 3.20, methionine+cystine 5.59 and phenylalanine+tyrosine 7.92. 2-Organoleptic evaluation revealed that shamy bread and biscuit samples fortified with 4% and 5% silver carp protein hydrolyzate, respectively were the best among all examined treatments; hence both contained more protein and essential amino acids contents

Histopathologic evaluation of the effect of microwave irradiation on fixation of kidney tissue

Post-mortem structural changes in tissues cause technical difficulties in accurate histo-pathologic interpretation. In the present study, we evaluated the effects of microwave irradiation before tissue fixation on preventing post mortem autolysis and accelerating the process of tissue fixation in the kidneys. Twenty-four chickens randomly were divided into 8 groups of 3 each. They were slaughtered and stored at room temperature. At 0, 1, 3, 5, 10, 15, 20 and 25 hours the kidneys were removed from chickens. The right kidneys were used as controls and fixed by conventional method through immersing in 10% buffered formalin; all the left kidneys were first irradiated by microwave [MWI; 30sec/450 watt] and then immersed in 10% buffered formalin. Kidney tissue samples were prepared according to standard routines, sliced into sections measuring 5 microns and stained by hematoxylin and eosin; next, these slides were examined through a light microscope. Morphologic study showed no difference in tissue preservation between the two methods till 20 hours post-mortem; however, at 25 hours, samples that had been fixed according to the standard methods revealed marked post-mortem structural degeneration while samples receiving microwave irradiation before fixation had preserved their original structure to a greater degree. Microwave irradiation before fixation of kidney tissue that has been dead for up to 25 hours results in greater structural preservation as compared to fixation by routine methods. We recommend further studies, preferably by using image analyzing, immunohistopathology and electronic microscopy to scrutinize minute structural changes in detail

Acute necrotizing pancreatitis and postmortem autolysis of pancreas / 法医学杂志

OBJECTIVE@#To compare the pathomorphologic changes between the pancreas in acute necrotizing pancreatitis (ANP) and that in acute deaths of rats (within 48 hours) so as to find the distinctions.@*METHODS@#The animal models of ANP and other acute deaths (electroshock, mechanic asphyxia/strangle, and acute poisoning with tetramine) were established according to the criteria. Half-quantitative grading and image quantitative analysis methods were employed to observe the gross and microscopic changes of the pancreases.@*RESULTS@#Three features including inflammation infiltrate, fat necrosis and calcium deposit in the ANP group were considerably different from that in other acutely died rat group (P<0.05).@*CONCLUSION@#Inflammation infiltrate, fat necrosis and calcium deposit are the most important pathologic features found in ANP by common light microscope, distinguishing ANP from postmortem pancreatic autolysis.

Indol-3-Carbinol Regulated Tight Junction Permeability and Associated-Protein Level and Suppressed Cell Invasion in Human Colon Cancer Cell Line, HT-29

To determine whether indol-3-carbinol (I3C, C9H9NO), an autolysis product of a glucosinolate and a glucobrassicin in vegetables, regulated tight junction proteins (TJ) and suppressed cell invasion in colon cancer cells,this experiment was performed. Our results indicate that I3C inhibit cell growth of HT-29 cells in a dose (0, 50, 100 micrometer) and time (0, 24 and 48 h) dependent manner. Using the wound healing and matrigel invasion study, respectively, I3C inhibits the cell motility and invasion of the ovarian cancer cell line. The TEER values were increased in HT-29 cells grown in transwells treated with I3C, reversely, paracellular permeability was decreased in those of condition. Claudin-1, claudin-5, ZO-1 and occuldin have been shown to be positively expressed in HT-29 coloncancer cells. I3C occurs concurrently with a significant decrease in the levels of those of proteins in HT-29 cells. But E-cadherin level in the HT-29 was increased by I3C. The reduction of claudin-1 and claudin-5 protein levels occurred post-transcriptionaly since their mRNA levels are no difference by I3C. Therefore, our results suggest that I3C may be expected to inhibit cancer metastasis and invasion by tighten the cell junction and restoring tight junction in colon cancer cell line, HT-29.

Effect of Indole-3-Carbinol on Inhibition of MMP Activity via MAPK Signaling Pathway in Human Prostate Cancer Cell Line, PC3 Cells

We examined the effect of indole-3-carbinol (I3C, C9H9NO), an autolysis product of a glucosinolate and a glucobrassicin in vegetables, on MMP-2, -9 activities and TIMP-1 and -2 inductions via microtubule-associated protein kinase (MAPK) signaling pathway in prostate cancer cell line, PC3 cells. Our results indicated that I3C inhibited cell growth of PC3 cells in dose (0, 50, 100 micrometer) and time (0, 24, 48 and 72 h) dependent manners. Using gelatin zymography for MMP activity, we demonstrated that I3C significantly decrease MMP-2 and -9 activities in PC3 cells. We also observed that I3C decreased the proteins and mRNA levels of MMP-2 and -9 in PC3 cells as well. Inversely, expressions of TIMP-1 and -2 protein and mRNA in PC3 cells were increased by I3C in a dose dependent manner. In another experiment, we showed that I3C inhibited PC3 cells invasiveness by using marigel invasion assay and we also found that I3C suppressed MMP transcriptional activity by MAPK signaling pathways. Taken together, our results suggest that I3C may contribute to the potential beneficial food component to prevent the cancer metastasis in prostate cancer cells.