Suitability of three different cereal grains for spawn development and their impact on the growth and yield of Macrocybe gigantea (Massee) Pegler & Lod.

Mushroom cultivation is an economical biotechnological process for the conversion of various unused lignocellulosic wastes into protein rich food. The present study was conducted to assess the suitability of three different cereal grains viz., bajra (Pennisetum glaucum L.), wheat (Triticum aestivum L.)  and maize (Zea mays L.) for spawn production of Macrocybe gigantea  (Massee) Pegler & Lod. and further its cultivation on two agrowastes (wheat straw and paddy straw) for assessing biological efficacy. It was observed that among the three cereal grains tested, bajra grains took significantly (P?0.05) less time for spawn development. Moreover, a minimum period of spawn run (16.3 days), highest sporophore yield (343.6g/500g of dry substrate) and biological efficiency (68.7%) were also recorded from substrate bags inoculated with bajra grain spawn. These results suggest the use of bajra grain spawn for quick and successful cultivation of M. gigantea

A randomized comparative study of exteriorization of uterus versus in situ intra-peritoneal repair at cesarean delivery

Background: Various caesarean delivery techniques have been compared in the past to assess the associated short-term and long-term advantages and disadvantages. Although uterine exteriorization at caesarean section is popular among obstetricians, safety of this technique remains a disputed matter. The aim of the present study was to compare the influence of uterine exteriorization or in situ repair on caesarean section morbidity.Methods: In this prospective, randomized, controlled study, 200 pregnant women with indication for caesarean delivery were randomized as 100 patients each in the exteriorization group and in the in-situ group. Data on mean time taken for uterine incision closure, intra-operative blood loss and post-operative morbidities were collected and compared between the two groups for statistical analysis.Results: A statistically significant trend towards lesser mean time taken for the uterine wound repair was observed in the exteriorization group. However significantly more number of patients had increased post-operative pain and need for additional analgesia in exteriorization group. There was no significant difference with respect to intra-operative blood loss and incidence of nausea and vomiting; incidence of post-operative endomyometritis, febrile morbidity, wound infection, time taken for return of bowel function and length of hospital stay among the two groups.Conclusions: We concluded that uterine exteriorization and in situ repair have similar post-operative caesarean section morbidity outcomes. However, in situ repair of uterus was associated with lesser post-operative pain, and exteriorization of uterus was associated with lesser operating time

Neutrophils instruct homeostatic and pathological states in naive tissues

Immune protection relies on the capacity of neutrophils to infiltrate challenged tissues. Naive tissues, in contrast, are believed to remain free of these cells and protected from their toxic cargo. Here, we show that neutrophils are endowed with the capacity to infiltrate multiple tissues in the steady-state, a process that follows tissue-specific dynamics. By focusing in two particular tissues, the intestine and the lungs, we find that neutrophils infiltrating the intestine are engulfed by resident macrophages, resulting in repression of Il23 transcription, reduced G-CSF in plasma, and reinforced activity of distant bone marrow niches. In contrast, diurnal accumulation of neutrophils within the pulmonary vasculature influenced circadian transcription in the lungs. Neutrophil-influenced transcripts in this organ were associated with carcinogenesis and migration. Consistently, we found that neutrophils dictated the diurnal patterns of lung invasion by melanoma cells. Homeostatic infiltration of tissues unveils a facet of neutrophil biology that supports organ function, but can also instigate pathological states.S

Neutrophil mobilization via plerixafor-mediated CXCR4 inhibition arises from lung demargination and blockade of neutrophil homing to the bone marrow

Blood neutrophil homeostasis is essential for successful host defense against invading pathogens. Circulating neutrophil counts are positively regulated by CXCR2 signaling and negatively regulated by the CXCR4-CXCL12 axis. In particular, G-CSF, a known CXCR2 signaler, and plerixafor, a CXCR4 antagonist, have both been shown to correct neutropenia in human patients. G-CSF directly induces neutrophil mobilization from the bone marrow (BM) into the blood, but the mechanisms underlying plerixafor-induced neutrophilia remain poorly defined. Using a combination of intravital multiphoton microscopy, genetically modified mice and novel in vivo homing assays, we demonstrate that G-CSF and plerixafor work through distinct mechanisms. In contrast to G-CSF, CXCR4 inhibition via plerixafor does not result in neutrophil mobilization from the BM. Instead, plerixafor augments the frequency of circulating neutrophils through their release from the marginated pool present in the lung, while simultaneously preventing neutrophil return to the BM. Our study demonstrates for the first time that drastic changes in blood neutrophils can originate from alternative reservoirs other than the BM, while implicating a role for CXCR4-CXCL12 interactions in regulating lung neutrophil margination. Collectively, our data provides valuable insights into the fundamental regulation of neutrophil homeostasis, which may lead to the development of improved treatment regimens for neutropenic patients.This research was funded by SIgN, A*STAR, Singapore. C.N.Z. Mattar and J.K.Y. Chan received salary support from the National Medical Research Council of Singapore (NMRC/TA/003/2012 and NMRC/CSA/012/2009, respectively).S

CXCR4 identifies transitional bone marrow premonocytes that replenish the mature monocyte pool for peripheral responses

It is well established that Ly6C(hi) monocytes develop from common monocyte progenitors (cMoPs) and reside in the bone marrow (BM) until they are mobilized into the circulation. In our study, we found that BM Ly6C(hi) monocytes are not a homogenous population, as current data would suggest. Using computational analysis approaches to interpret multidimensional datasets, we demonstrate that BM Ly6C(hi) monocytes consist of two distinct subpopulations (CXCR4(hi) and CXCR4(lo) subpopulations) in both mice and humans. Transcriptome studies and in vivo assays revealed functional differences between the two subpopulations. Notably, the CXCR4(hi) subset proliferates and is immobilized in the BM for the replenishment of functionally mature CXCR4(lo) monocytes. We propose that the CXCR4(hi) subset represents a transitional premonocyte population, and that this sequential step of maturation from cMoPs serves to maintain a stable pool of BM monocytes. Additionally, reduced CXCR4 expression on monocytes, upon their exit into the circulation, does not reflect its diminished role in monocyte biology. Specifically, CXCR4 regulates monocyte peripheral cellular activities by governing their circadian oscillations and pulmonary margination, which contributes toward lung injury and sepsis mortality. Together, our study demonstrates the multifaceted role of CXCR4 in defining BM monocyte heterogeneity and in regulating their function in peripheral tissues

Mechanisms of glomerular leukocyte trafficking

Multiple forms of glomerulonephritis (GN) result from inappropriate recruitment of leukocytes into glomeruli. As such, it is essential that the mechanisms driving leukocyte recruitment to the glomerular vasculature are understood. Using an acute model of GN induced by an antibody against the glomerular basement membrane (GBM), it was previously demonstrated that glomerular leukocyte recruitment is dependent on the initial accumulation of platelets to the glomerulus. However, the molecular mechanisms of platelet recruitment to the glomerulus under these conditions are unknown. In addition, with the use of intravital microscopy, studies have demonstrated that leukocyte recruitment to the inflamed glomerulus can occur without an initial rolling interaction normally observed in post-capillary venules. In contrast, leukocytes undergo immediate arrest within glomerular capillaries. However, the behavior of neutrophils after this initial arrest interaction is poorly understood. Multiphoton confocal microscopy (MPCM) is a form of in vivo imaging which allows continuous visualization of leukocyte behavior over extended periods. Therefore the aims of the work described in this thesis were to further our understanding of the interactions required for induction of leukocyte recruitment upon glomerular inflammation. To achieve these aims, mice were treated with anti-GBM antibody and glomeruli were visualized using in vivo imaging techniques. The first set of studies examined the molecular basis of glomerular platelet recruitment. Platelets were isolated from a donor mouse, labeled with a fluorochrome then infused into a recipient mouse during conventional intravital microscopy experiments. Using this approach, it was observed that platelets lacking the platelet collagen receptor GPVI were reduced in their ability to undergo glomerular platelet recruitment. Similarly, inhibition of the platelet integrin αIIbβ3 or fibrinogen caused significant reductions in anti-GBM antibody-induced platelet adhesion. Using immunohistochemistry, fibrinogen deposition was detected in inflamed glomeruli, occurring via a partially ICAM-1-dependent pathway. Studies in neutrophil-depleted mice undergoing glomerular inflammation demonstrated a role for neutrophils in glomerular platelet accumulation. In contrast, inhibition of adenosine diphosphate-dependent platelet activation did not affect platelet recruitment to the inflamed glomerulus. Taken together, these findings indicate that the combined actions of GPVI and the αIIbβ3/fibrinogen/ICAM-1 pathway contribute to platelet recruitment in the inflamed glomerulus. The aim of the second set of studies was to use in vivo MPCM to investigate leukocyte recruitment to the inflamed glomerulus. This was achieved by visualizing glomerular capillaries and neutrophils by labeling them with specific fluorochromes. Under control conditions, neutrophils were observed to adhere and persist in the glomerular capillaries for an average of 5 minutes (‘dwell time’), with some of these cells remaining static for this period, and others undergoing migration within the vasculature. In contrast, in response to anti-GBM antibody, neutrophil dwell time was significantly increased in the first two hours post anti-GBM antibody treatment, with this change applying to both static and migratory cells. This study extends our understanding of neutrophil behavior following recruitment to the inflamed glomerulus showing that induction of glomerular inflammation does not alter the number of leukocyte recruited to the glomerulus, but increases the duration of their retention. The aim of the next series of experiments was to utilize the MPCM imaging approach to characterize the mechanisms of glomerular leukocyte recruitment. Inhibition of the β2 integrin significantly reduced the dwell time of migrating neutrophils in the first hour of the anti-GBM antibody response, whereas during the second hour, inhibition of the β2 integrin Mac-1 significantly reduced neutrophil dwell time for both static and migratory cells. Anti-Mac-1 treatment also significantly decreased urinary protein excretion, indicating that Mac-1 is a key leukocyte integrin responsible for leukocyte recruitment to inflamed glomerular capillaries. Finally, direct examination of neutrophils during glomerular inflammation in vivo demonstrated that both static and crawling neutrophils generated oxidants in inflamed glomeruli. In summary, these experiments demonstrate that the process of glomerular leukocyte recruitment is more dynamic than previously recognized. Moreover, the application of new imaging technique has revealed novel information about neutrophil behavior in the glomerular vasculature

The genetics of dilated cardiomyopathy

Dilated cardiomyopathy (DCM) is a leading cause for cardiac transplantation with an estimated prevalence of 36.5 per 100,000 individuals in the USA.  DCM is a disorder characterized by ventricular dilation and systolic contractile dysfunction.  It is the result of a heterogenous group of inherited and acquired disorders and is an important cause of heart failure.  Idiopathic DCM is the most common cause of DCM and accounts for ~50% of cases with a familial basis found in up to 35%. A genome wide screen in four generations of a Puerto Rican kindred with autosomal dominant isolated familial dilated cardiomyopathy (FDCM) was conducted.  Two-point linkage analysis mapped the disease locus to chromosome 14q11.2-13 (lod score = 5.11 at q  = 0).  The b-myosin heavy chain (b-MHC) candidate gene mapped to this region and direct sequencing identified a single nucleotide T168C mutation (Ser®Pro532) which was highly conserved across a range of different species.  A novel mismatch primer that converted the half-site to a full recognition site for the SphI endonuclease independently confirmed the mutation and its absence in 192 normal chromosomes.  A further disease causing mutation 2378C ®G  replacing Phe764Leu in b-MHC was identified in an unrelated family.  A further two unrelated FDCM families were found to have deleted one of four tandem lysine residues (DLys210) of the cardiac troponin T gene. Understanding the monogenic disorders provides us with important knowledge not only about the rare condition being studied but also about potential mechanisms underlying the more common phenotypes.  This finding may open up novel and perhaps unpredictable therapeutic targets.EThOS - Electronic Theses Online ServiceGBUnited Kingdo

Mechanisms of glomerular leukocyte trafficking

Multiple forms of glomerulonephritis (GN) result from inappropriate recruitment of leukocytes into glomeruli. As such, it is essential that the mechanisms driving leukocyte recruitment to the glomerular vasculature are understood. Using an acute model of GN induced by an antibody against the glomerular basement membrane (GBM), it was previously demonstrated that glomerular leukocyte recruitment is dependent on the initial accumulation of platelets to the glomerulus. However, the molecular mechanisms of platelet recruitment to the glomerulus under these conditions are unknown. In addition, with the use of intravital microscopy, studies have demonstrated that leukocyte recruitment to the inflamed glomerulus can occur without an initial rolling interaction normally observed in post-capillary venules. In contrast, leukocytes undergo immediate arrest within glomerular capillaries. However, the behavior of neutrophils after this initial arrest interaction is poorly understood. Multiphoton confocal microscopy (MPCM) is a form of in vivo imaging which allows continuous visualization of leukocyte behavior over extended periods. Therefore the aims of the work described in this thesis were to further our understanding of the interactions required for induction of leukocyte recruitment upon glomerular inflammation. To achieve these aims, mice were treated with anti-GBM antibody and glomeruli were visualized using in vivo imaging techniques. The first set of studies examined the molecular basis of glomerular platelet recruitment. Platelets were isolated from a donor mouse, labeled with a fluorochrome then infused into a recipient mouse during conventional intravital microscopy experiments. Using this approach, it was observed that platelets lacking the platelet collagen receptor GPVI were reduced in their ability to undergo glomerular platelet recruitment. Similarly, inhibition of the platelet integrin αIIbβ3 or fibrinogen caused significant reductions in anti-GBM antibody-induced platelet adhesion. Using immunohistochemistry, fibrinogen deposition was detected in inflamed glomeruli, occurring via a partially ICAM-1-dependent pathway. Studies in neutrophil-depleted mice undergoing glomerular inflammation demonstrated a role for neutrophils in glomerular platelet accumulation. In contrast, inhibition of adenosine diphosphate-dependent platelet activation did not affect platelet recruitment to the inflamed glomerulus. Taken together, these findings indicate that the combined actions of GPVI and the αIIbβ3/fibrinogen/ICAM-1 pathway contribute to platelet recruitment in the inflamed glomerulus. The aim of the second set of studies was to use in vivo MPCM to investigate leukocyte recruitment to the inflamed glomerulus. This was achieved by visualizing glomerular capillaries and neutrophils by labeling them with specific fluorochromes. Under control conditions, neutrophils were observed to adhere and persist in the glomerular capillaries for an average of 5 minutes (‘dwell time’), with some of these cells remaining static for this period, and others undergoing migration within the vasculature. In contrast, in response to anti-GBM antibody, neutrophil dwell time was significantly increased in the first two hours post anti-GBM antibody treatment, with this change applying to both static and migratory cells. This study extends our understanding of neutrophil behavior following recruitment to the inflamed glomerulus showing that induction of glomerular inflammation does not alter the number of leukocyte recruited to the glomerulus, but increases the duration of their retention. The aim of the next series of experiments was to utilize the MPCM imaging approach to characterize the mechanisms of glomerular leukocyte recruitment. Inhibition of the β2 integrin significantly reduced the dwell time of migrating neutrophils in the first hour of the anti-GBM antibody response, whereas during the second hour, inhibition of the β2 integrin Mac-1 significantly reduced neutrophil dwell time for both static and migratory cells. Anti-Mac-1 treatment also significantly decreased urinary protein excretion, indicating that Mac-1 is a key leukocyte integrin responsible for leukocyte recruitment to inflamed glomerular capillaries. Finally, direct examination of neutrophils during glomerular inflammation in vivo demonstrated that both static and crawling neutrophils generated oxidants in inflamed glomeruli. In summary, these experiments demonstrate that the process of glomerular leukocyte recruitment is more dynamic than previously recognized. Moreover, the application of new imaging technique has revealed novel information about neutrophil behavior in the glomerular vasculature

A randomized comparative study of exteriorization of uterus versus in situ intra-peritoneal repair at cesarean delivery

Background: Various caesarean delivery techniques have been compared in the past to assess the associated short-term and long-term advantages and disadvantages. Although uterine exteriorization at caesarean section is popular among obstetricians, safety of this technique remains a disputed matter. The aim of the present study was to compare the influence of uterine exteriorization or in situ repair on caesarean section morbidity.Methods: In this prospective, randomized, controlled study, 200 pregnant women with indication for caesarean delivery were randomized as 100 patients each in the exteriorization group and in the in-situ group. Data on mean time taken for uterine incision closure, intra-operative blood loss and post-operative morbidities were collected and compared between the two groups for statistical analysis.Results: A statistically significant trend towards lesser mean time taken for the uterine wound repair was observed in the exteriorization group. However significantly more number of patients had increased post-operative pain and need for additional analgesia in exteriorization group. There was no significant difference with respect to intra-operative blood loss and incidence of nausea and vomiting; incidence of post-operative endomyometritis, febrile morbidity, wound infection, time taken for return of bowel function and length of hospital stay among the two groups.Conclusions: We concluded that uterine exteriorization and in situ repair have similar post-operative caesarean section morbidity outcomes. However, in situ repair of uterus was associated with lesser post-operative pain, and exteriorization of uterus was associated with lesser operating time

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