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Biomedical subjects

R Taneja

Publications and source records attributed to R Taneja.

At least 19 recordsLinked to original sources
PubMed2002

Surgeon workload and motion efficiency with robot and human laparoscopic camera control.

BACKGROUND: Surgeons are now being assisted by robotic systems in a wide range of laparoscopic procedures. Some reports have suggested that robot-assisted camera control (RACC) may be superior to a human driver in terms of quality of view and directional precision, as well as long-term cost savings. Therefore, we setout to investigate the impact of RACC of surgeon motion efficiency. METHODS: Twenty pigs were randomized to undergo a standardized laparoscopic Nissen fundoplication with either a human or RACC system, the AESOP 2000. All procedures were performed by the same surgical fellow. Time was recorded for dissection and suture phases. Inertial motion sensors were used to monitor both the surgeon's hands and the camera. Digitized data were analyzed to produce summary measures related to overall motion. RESULTS: The operative times were slightly longer with RACC (mean 80.2 +/- 20.6 vs 73.1 +/- 15.4 min, not significant). With regard to operative times and surgeon motion measures, the only statistically significant differences were for setup and breakdown times, which contributed <15% to the total time for the procedure. CONCLUSION: In terms of impact on surgeon motion efficiency and operative time under normal surgical conditions, RACC is essentially the same as an expert human driver. However, careful planning and structuring of the surgical suite may yield some small gains in operative time.

Animals↗
PubMed2001

Cross-regulatory interaction between Stra13 and USF results in functional antagonism.

Transcription factors belonging to the basic helix-loop-helix (bHLH) family are critical regulators of cellular proliferation and differentiation. The functional activity of these proteins can be regulated by heterodimerization through the HLH domain, as a result of formation of functional or non-functional heterodimers. The presence of a leucine zipper in bHLH-leucine zipper (bHLHZip) proteins, however, prevents such heterodimeric interactions via the HLH domain between bHLH and bHLHZip proteins. To identify cellular proteins that directly interact with and modulate transcriptional repression mediated by the bHLH protein Stra13, we carried out a yeast two hybrid screen. The bHLHZip protein USF (Upstream Stimulatory factor) was identified as a Stra13 interacting protein. We demonstrate a direct interaction between Stra13 and USF that is dependent upon the C-terminal repression domain of Stra13 and the DNA-binding domain of USF. Stra13 and USF also colocalize and functionally interact in mammalian cells. Co-expression of USF abrogates Stra13-mediated repression of target genes and conversely, Stra13 inhibits DNA-binding and USF-mediated transactivation. Taken together, our data demonstrate that Stra13 and USF interact physically and functionally, and identify a novel mode of cross regulatory interaction between members of the bHLH and bHLHZip families that abrogates their functional activity.

Animals↗
PubMed2001

Targeted drug delivery systems 6: Intracellular bioreductive activation, uptake and transport of an anticancer drug delivery system across intestinal Caco-2 cell monolayers.

We demonstrate transport across, intracellular accumulation and bioreductive activation of a conformationally constrained, anticancer drug delivery system (the CH(3)-TDDS) using Caco-2 cell monolayers (CCMs) as an in vitro model of the human intestinal mucosa. Reverse-phase High Performance Liquid Chromatography (HPLC) coupled with UV detection was used to detect CH(3)-TDDS, the bioreduction product (lactone) and the released drug (melphalan methyl ester; MME). Upon incubation of the CH(3)-TDDS with the apical (AP) surface of 21-day-old CCM, we observed rapid decrease in the AP concentration of the CH(3)-TDDS (60%/hr) as a result of cellular uptake. Rapid intracellular accumulation of the CH(3)-TDDS was followed by bioreductive activation to deplete the cellular levels of CH(3)-TDDS. The drug part (MME) and lactone, as well as CH(3)-TDDS, were detected in the basolateral (BL) chamber. Intracellular Caco-2 levels of TDDS and lactone were also detectable. Bioreductive activation of the CH(3)-TDDS was additionally confirmed by formation of lactone after incubation of the CH(3)-TDDS in the presence of freshly prepared Caco-2 cell homogenates. During transport studies of melphalan or MME alone (as control), the intact drug was not detected in the intracellular compartment or in the BL chamber. These observations demonstrate that CH(3)-TDDS has potential for improving intestinal delivery of MME. TDDS could be useful in facilitating oral absorption of MME as well as the oral delivery of other agents.

Antineoplastic Agents↗
PubMed2001

Defective T cell activation and autoimmune disorder in Stra13-deficient mice.

Stra13, a basic helix-loop-helix transcription factor, is up-regulated upon activation of CD4+ T cells. Here we show that Stra13-deficient mice exhibit defects in several phases of CD4+ T cell activation. In vivo, Stra13 deficiency results in ineffective elimination of activated T and B cells, which accumulate progressively, leading to lymphoid organ hyperplasia. Consequently, aging Stra13-/- mice develop autoimmune disease characterized by accumulation of spontaneously activated T and B cells, circulating autoantibodies, infiltration of T and B lymphocytes in several organs and immune complex deposition in glomeruli. Our studies identify Stra13 as a key regulator of lymphocyte activation that is vital for maintenance of self-tolerance and for constraint of autoimmunity.

Aging↗
PubMed2000

Stra13 expression is associated with growth arrest and represses transcription through histone deacetylase (HDAC)-dependent and HDAC-independent mechanisms.

Stra13 is a transcriptional repressor related within its basic helix-loop-helix domain with the Drosophila Hairy, Enhancer of Split, and the mouse Hes1 proteins that interact with the corepressor Groucho. Because Stra13 lacks the conserved WRPW motif for interaction with Groucho, we examined the function and mechanism of transcriptional repression mediated by Stra13 that exhibits several distinctive features. Here, we report that Stra13 expression is closely associated with cell growth arrest induced by several triggers such as retinoic acid and trichostatin A (TSA; a specific histone deacetylase inhibitor) as well as by serum starvation. Stra13 expression is transcriptionally repressed and maintained at a low level in cells through a negative autoregulatory mechanism that is brought about by its interaction with the corepressor histone deacetylase (HDAC1). This interaction requires the Stra13 C-terminal domain containing three alpha-helices, which are also functionally critical to its repressive activity. Thus, inhibition of HDAC activity by TSA abrogates Stra13-mediated repression of its promoter, resulting in induction of Stra13 expression that is coincident with TSA-induced growth arrest. Further, once induced, Stra13 strongly represses the expression of the cell proliferation-associated gene c-Myc through an HDAC1-independent pathway that involves its interaction with the basal transcription factor TFIIB. Our studies indicate that Stra13 may play a key role in signaling pathways that lead to growth arrest and terminal differentiation by repression of target genes via HDAC-dependent and HDAC-independent mechanisms.

3T3 Cells↗
PubMed2000

Effects of hypoxia on granulocytic-monocytic progenitors in rats. Role of bone marrow stroma.

Hemorrhagic shock leads to hypoxia and is associated with bone marrow (BM) failure. Hemorrhagic shock is also a predisposing factor in immune dysregulation. Since the BM is the major organ of immune cells in the adult, its failure following hemorrhagic shock may explain the increased susceptibility to infection. The in vitro evidence indicates that hypoxia mediates altered functions in BM stroma. Since similar hematopoietic alterations are reported in hypoxia and hemorrhagic shock, hypoxia alone could be a representative model to study BM responses during hemorrhagic shock. In this study, we use an animal model to dissect the hematopoietic effects of hypoxia. We subjected rats to hypoxia, and at days 1 and 5 post-hypoxia we determined the numbers of granulocytic-monocytic progenitors (CFU-GM) in the BM. We found significant increase (P < 0.05) in CFU-GM at day 1 and a downward trend by day 5. Enhanced BM cellularity could not explain the increase in CFU-GM by day 1. BM stromal cells mediated most of the stimulatory effects by hypoxia. CFU-GM was inversely proportional to bioactive TGF-beta and directly proportional to IL-1. Compared to normoxic rats, IL-6 production was suppressed in BM cells from hypoxic rats. The results show that hypoxia alone initiate a stimulatory response in CFU-GM progenitors. These effects are at least partially mediated through the BM stroma. In the absence of a second insult, CFU-GM reverts to baseline. The data also suggest that hypoxia mediates complex responses that include cytokine production. These results add to the current understanding of hematopoietic responses by hypoxia and adds to the mechanisms of immune dysfunctions following hemorrhagic shock.

Animals↗
PubMed2000

Phagocytosis of Candida albicans induces apoptosis of human neutrophils.

Neutrophil-mediated inflammation is terminated through the programmed cell death or apoptosis of the neutrophil, a process that can be inhibited by soluble mediators released during an inflammatory response. It has been reported, however, that the phagocytosis of intact bacteria can accelerate apoptosis. We evaluated the effects of the phagocytosis of a common nosocomial pathogen, Candida albicans, on the expression of apoptosis. Phagocytosis of killed Candida induced a dose-dependent increase in the apoptosis of normal neutrophils after 18 h of in vitro culture, from 40.7+/-9.1% to 81.7+/-4.5%, while supernatants from neutrophil:Candida co-cultures actually inhibited apoptosis. Induction of apoptosis was not dependent on phagocytosis, since opsonization of yeast with serum failed to increase apoptosis, while inhibition of phagocytosis with latrunculin B resulted in a slightly increased apoptotic rate. Increased apoptosis induced by Candida was associated with increased activity of the membrane-associated apoptotic enzyme, caspase 8, and with increased expression of the active form of the key executioner caspase, caspase 3. Increased apoptosis was associated with depletion of intracellular glutathione (GSH), and could be inhibited by the addition of exogenous GSH. These data demonstrate an important physiologic role for host-pathogen interactions in the resolution of inflammation and suggest that the response to an invading pathogen is an important stimulus to the restoration of normal immunologic homeostasis.

Apoptosis↗
PubMed2000

The AF-1 and AF-2 activating domains of retinoic acid receptor-alpha (RARalpha) and their phosphorylation are differentially involved in parietal endodermal differentiation of F9 cells and retinoid-induced expression of target genes.

Retinoic acid (RA) induces the differentiation of F9 cells cultured as monolayers into primitive endodermal-like cells, whereas a combination of RA and cAMP leads to parietal endodermal differentiation. In RA receptor alpha-null F9 cells (RARalpha-/- cells), RA still efficiently triggers RARgamma-mediated primitive endodermal differentiation, but parietal endodermal differentiation is markedly delayed. To investigate the role of RARalpha1 activation functions AF-1 and AF-2 and of their phosphorylation sites during RA- and cAMP-induced parietal differentiation, cell lines reexpressing WT or mutated RARalpha1 were established in RARalpha-/- cells. We have found that the protein kinase A (PKA) phosphorylation site and the AF-2AD core (helix 12) of RARalpha1 are required for efficient parietal endodermal differentiation, whereas the AF-1 proline-directed kinase phosphorylation site is dispensible. Interestingly, deletion of the AF-1 activating domain (the A/B region), but not of the AF-2AD core, generates a dominant negative mutant that abrogates primitive endodermal differentiation when expressed in RARalpha-/- cells. We also show that the RARalpha AF-1 and AF-2 activation functions, but not their phosphorylation sites, are involved in the induction of RA-responsive genes in a differential promoter context-dependent manner.

Animals↗
PubMed1999

Prediction of dissolution-absorption relationships from a continuous dissolution/Caco-2 system.

The objectives were 1) to design a continuous dissolution/Caco-2 system to predict the dissolution-absorption relationships for fast and slow dissolving formulations of piroxicam, metoprolol tartrate, and ranitidine HCl, and compare the predicted relationships with observed relationships from clinical studies; 2) to estimate the effect of croscarmellose sodium on ranitidine dissolution-absorption relationships; and 3) to estimate the effect of solubilizing agents on piroxicam dissolution-absorption relationships. A continuous dissolution/Caco-2 system was constructed from a dissolution apparatus and a diffusion cell, such that drug dissolution and permeation across a Caco-2 monolayer would occur sequentially and simultaneously. The continuous system generally matched observed dissolution-absorption relationships from clinical studies. For example, the system successfully predicted the slow metoprolol and slow ranitidiine formulations to be permeation-rate-limited. The system predicted the slow piroxicam formulation to be dissolution-rate-limited, and the fast piroxicam formulation to be permeation-rate-limited, in spite of piroxicam's high permeability and low solubility. Additionally, the system indicated croscarmellose sodium enhanced ranitidine permeability and predicted solubilizing agents to not modulate permeability. These results suggest a dissolution/Caco-2 system to be an experimentally based tool that may predict dissolution-absorption relationships from oral solid dosage forms, and hence the relative contributions of dissolution and permeation to oral drug absorption kinetics.

Absorption↗
PubMed1997

Phosphorylation of activation functions AF-1 and AF-2 of RAR alpha and RAR gamma is indispensable for differentiation of F9 cells upon retinoic acid and cAMP treatment.

The role of RAR alpha 1 and RAR gamma 2 AF-1 and AF-2 activation functions and of their phosphorylation was investigated during RA-induced primitive and parietal differentiation of F9 cells. We found that: (i) primitive endodermal differentiation requires RAR gamma 2, whereas parietal endodermal differentiation requires both RAR gamma 2 and RAR alpha 1, and in all cases AF-1 and AF-2 must synergize; (ii) primitive endodermal differentiation requires the proline-directed kinase site of RAR gamma 2-AF-1, whereas parietal endodermal differentiation additionally requires that of RAR alpha 1-AF-1; (iii) the cAMP-induced parietal endodermal differentiation also requires the protein kinase A site of RAR alpha-AF-2, but not that of RAR gamma; and (iv) the AF-1-AF-2 synergism and AF-1 phosphorylation site requirements for RA-responsive gene induction are promoter context-dependent. Thus, AF-1 and AF-2 of distinct RARs exert specific cellular and molecular functions in a cell-autonomous system mimicking physiological situations, and their phosphorylation by kinases belonging to two main signalling pathways is required to enable RARs to transduce the RA signal during F9 cell differentiation.

Animals↗
PubMed1997

Overexpression of Stra13, a novel retinoic acid-inducible gene of the basic helix-loop-helix family, inhibits mesodermal and promotes neuronal differentiation of P19 cells.

We report the cDNA cloning of Stra13, a novel retinoic acid (RA)-inducible gene from P19 embryonal carcinoma cells that encodes a basic helix-loop-helix (bHLH) protein that shows the highest sequence similarities to the Drosophila Hairy and Enhancer of split and mouse Hes proteins. Stra13 does not bind to the known consensus motifs (E-box and N-box) for bHLH proteins, but can repress activated transcription (through an alpha-helix rich domain) in part by interaction with general factors of the basal transcription machinery. During mouse embryogenesis, Stra13 RNA is expressed in the neuroectoderm, and also in a number of mesodermal and endodermal derivatives. Remarkably, overexpression of Stra13 in P19 cells results in neuronal differentiation in monolayer culture, under conditions where wild-type P19 cells only undergo mesodermal/endodermal differentiation. This neuronal differentiation is accompanied by an altered expression of mesodermal and neuronal markers, indicating that Stra13 could be one of the earliest RA target genes whose expression is required for repression of mesodermal/endodermal differentiation and/or induction of neuronal differentiation when P19 cell aggregates are exposed to RA. Our results raise the possibility that Stra13 could be involved as a repressor in a number of decision events occurring during differentiation of various cell lineages.

Amino Acid Sequence↗
PubMed1997

Differential effects of acute hypoxia and endotoxin on the secretion and expression of bone marrow interleukin-1 and interleukin-6.

Hemorrhagic shock induces tissue hypoxia and has been demonstrated to alter the myelopoietic response to bacterial lipopolysaccharide (LPS). Interleukin-1 and interleukin-6 are important mediators of immunologic events after hemorrhagic shock. Bone marrow stroma release inflammatory cytokines, which may play a role in the regulation of myelopoiesis after injury. The aim of this study was to correlate cytokine gene expression with protein release and myelopoiesis by total bone marrow cells. The role of bone marrow stroma after exposure to hypoxia and lipopolysaccharide was also examined. BALB/c mice were designated as normoxia or hypoxia and total bone marrow cells were harvested. Hypoxia mice were exposed to 2 h of 5% O2/95% N2, and then returned to room air. Additional groups of mice were given LPS intraperitoneally. Bone marrow stroma, from BALB/c mice, was similarly designated. Myelopoiesis was assessed by growth of granulocyte-macrophage progenitor cells (CFU-GM). Interleukin-1 and interleukin-6 protein activity was assessed by bioassay. RNA was extracted from both total bone marrow cells and bone marrow stroma. By day 5, LPS alone resulted in a 93% increase in CFU-GM versus normoxia. Hypoxia and LPS exposure significantly decreased CFU-GM on days 1, 3, and 5. LPS alone induced an increase in interleukin-6. At 2, 6, and 24 h, hypoxia blunted interleukin-6 release in response to LPS. Hypoxia alone could not induce interleukin-6. However, hypoxia did induce interleukin-1 mRNA without the release of bioactive protein. In the remainder of groups, interleukin-1 protein levels and mRNA levels were correlated. Bone marrow stroma interleukin-1 and interleukin-6 protein activity was consistently correlated with that of total bone marrow. These data demonstrate that bone marrow cytokine production is differentially regulated by hypoxia. Hypoxia impairs interleukin-6 protein and mRNA in response to LPS, which may play a role in the suppression of myelopoiesis after shock. Also, bone marrow stroma plays an integral role in regulating myelopoiesis.

Animals↗