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Biomedical subjects

S N Bhatia

Publications and source records attributed to S N Bhatia.

At least 19 recordsLinked to original sources

Distinct phenotypic consequences of cholangiocarcinoma-associated FGFR2 alterations depend on biliary epithelial cell state.

Epithelial cancers disrupt tissue architecture and are often driven by mutations in genes that play important roles in normal epithelial morphogenesis. The intrahepatic biliary system is an epithelial tubular network that forms within the developing liver via the de novo initiation and expansion of apical lumens. Intrahepatic biliary tumors (intrahepatic cholangiocarcinoma) commonly harbor activating genomic alterations in the FGFR2 receptor tyrosine kinase, which plays important roles in epithelial morphogenesis in other developmental settings. Using a physiologic and quantitative 3D model we demonstrate that FGFR signaling is important for biliary morphogenesis and that oncogenic FGFR2 fusions and in-frame deletions disrupt biliary architecture. Importantly, we show that the trafficking of and signaling from the FGFR2 mutants, as well as their phenotypic impacts, are governed by the epithelial state of the cell. Unexpectedly, we also found that distinct tumor-driving FGFR2 mutants disrupt biliary morphogenesis in completely different and clinically relevant ways, informing our understanding of morphogenesis and tumorigenesis and highlighting the importance of convergent studies of both.

Journal Article↗

Oxygen consumption characteristics of porcine hepatocytes.

Oxygen uptake rate (OUR) of hepatocytes is an important parameter for the design of bioartificial liver assist (BAL) devices. Porcine hepatocytes were cultured in a specially constructed measurement chamber with an incorporated mixing system and a Clark polarographic oxygen electrode. Signal noise associated with conventional Clark electrode implementations was circumvented by the combination of real time digital numerical averaging and subsequent finite impulse response (FIR) spectral filtering. Additional software allowed for the automated generation of cellular oxygen consumption coefficients, namely, Vmax and K0.5, adding a high degree of objectivity to parameter determination. Optimization of the above numerical techniques identified a 0.1 Hz/200 data point sample size and a 0.004 Hz cutoff frequency as ideal parameters. Vmax values obtained for porcine hepatocytes during the first two weeks of culture showed a maximal consumption of 0.9 nmole/sec/10(6) cells occurring on Day 4 post seeding, and a gradual decrease to 0.31 nmole/sec/10(6) cells by Day 15. K0.5 values increased from 2 mm Hg on Day 2 to 8 mm Hg by Day 8, with gradual subsequent decrease to 4 mm Hg by Day 15. The Vmax and K0.5 values measured for porcine cells were higher than maximal values for rat hepatocytes (Vmax: 0.43 nmole/sec/10(6) cells, K0.5: 5.6 mmHg) and thus may necessitate significantly altered BAL device design conditions to ensure no oxygen limitations. Finally, these results highlight the need for species specific characterization of cellular function for optimal BAL device implementations.

Animals↗

Effect of cell-cell interactions in preservation of cellular phenotype: cocultivation of hepatocytes and nonparenchymal cells.

Heterotypic cell interaction between parenchymal cells and nonparenchymal neighbors has been reported to modulate cell growth, migration, and/or differentiation. In both the developing and adult liver, cell-cell interactions are imperative for coordinated organ function. In vitro, cocultivation of hepatocytes and nonparenchymal cells has been used to preserve and modulate the hepatocyte phenotype. We summarize previous studies in this area as well as recent advances in microfabrication that have allowed for more precise control over cell-cell interactions through 'cellular patterning' or 'micropatterning'. Although the precise mechanisms by which nonparenchymal cells modulate the hepatocyte phenotype remain unelucidated, some new insights on the modes of cell signaling, the extent of cell-cell interaction, and the ratio of cell populations are noted. Proposed clinical applications of hepatocyte cocultures, typically extracorporeal bioartificial liver support systems, are reviewed in the context of these new findings. Continued advances in microfabrication and cell culture will allow further study of the role of cell communication in physiological and pathophysiological processes as well as in the development of functional tissue constructs for medical applications.

Animals↗

Numerical model of fluid flow and oxygen transport in a radial-flow microchannel containing hepatocytes.

The incorporation of monolayers of cultured hepatocytes into an extracorporeal perfusion system has become a promising approach for the development of a temporary bioartificial liver (BAL) support system. In this paper we present a numerical investigation of the oxygen tension, shear stress, and pressure drop in a bioreactor for a BAL composed of plasma-perfused chambers containing monolayers of porcine hepatocytes. The chambers consist of microfabricated parallel disks with center-to-edge radial flow. The oxygen uptake rate (OUR), measured in vitro for porcine hepatocytes, was curve-fitted using Michaelis-Menten kinetics for simulation of the oxygen concentration profile. The effect of different parameters that may influence the oxygen transport inside the chambers, such as the plasma flow rate, the chamber height, the initial oxygen tension in the perfused plasma, the OUR, and K(m) was investigated. We found that both the plasma flow rate and the initial oxygen tension may have an important effect upon oxygen transport. Increasing the flow rate and/or the inlet oxygen tension resulted in improved oxygen transport to cells in the radial-flow microchannels, and allowed significantly greater diameter reactor without oxygen limitation to the hepatocytes. In the range investigated in this paper (10 microns < H < 100 microns), and for a constant plasma flow rate, the chamber height, H, had a negligible effect on the oxygen transport to hepatocytes. On the contrary, it strongly affected the mechanical stress on the cells that is also crucial for the successful design of the BAL reactors. A twofold decrease in chamber height from 50 to 25 microns produced approximately a fivefold increase in maximal shear stress at the inlet of the reactor from 2 to 10 dyn/cm2. Further decrease in chamber height resulted in shear stress values that are physiologically unrealistic. Therefore, the channel height needs to be carefully chosen in a BAL design to avoid deleterious hydrodynamic effects on hepatocytes.

Animals↗

Prenatal detection and mapping of a distal 8p deletion associated with congenital heart disease.

We report the prenatal diagnosis, at 18 weeks' gestational age of a del(8)(p23.1-->pter) in a fetus with an atrio-ventricular canal, persistent left superior vena cava and hypoplastic right ventricle detected by sonographic imaging. We further refine the breakpoints associated with this defect using fluorescent in situ hybridization analysis (FISH). Our findings correlate with recent reports of the localization and importance of GATA4 (a zinc finger transcription factor) in cardiac development. Though microcephaly, mental retardation and typical behavioural features are well described in various deletions in 8p, the absence of notable microcephaly in this case raises the possibility for a separate genetic aetiology for some of these features. Indeed, primary autosomal recessive microcephaly (MCPH1) was recently mapped to a nearby region and may be the cause for this frequent observation in some cases of 8p deletions. These observations illustrate the role of FISH in prenatal diagnosis and refinement of chromosomal breakpoints. In addition, mappings of loci significant for cardiac development are presented. Our findings suggest that some features of the 8p deletion syndrome may ultimately be uncoupled from one another, and underscore the need for further study of this region of chromosome 8, in order to achieve adequate information for genetic counselling.

Adult↗

Probing heterotypic cell interactions: hepatocyte function in microfabricated co-cultures.

Replacement of liver function using extracorporeal bioartificial systems has been attempted with limited success. The instability of the hepatocyte phenotype in vitro has restricted the useful lifetime of these devices. Co-cultivation of hepatocytes with mesenchymal cells is one method that has been widely utilized to stabilize the liver-specific function of isolated cells; however, co-culture has yet to be successfully incorporated in a bioreactor setting. In this study, we probed heterotypic cell interactions in co-cultures of hepatocytes and 3T3 in order to better understand the cellular microenvironment necessary to induce and stabilize liver-specific functions. Using microfabrication and conventional techniques to control the heterotypic interface, the effects of varying degrees of heterotypic interaction on tissue function (albumin and urea synthesis) were examined. Our data indicated maximal induction of liver-specific functions in cultures with maximal initial heterotypic interaction, and that induction of hepatic functions in hepatocytes was increased in the vicinity of fibroblasts as compared to hepatocytes far from the heterotypic interface. Furthermore, our data suggested that heterotypic cell contact is necessary for induction of these functions. These studies will aid in the formation of design criteria for a co-culture based bioartificial liver, as well as provide a useful tool to study the role of heterotypic and homotypic interactions in liver physiology and pathophysiology.

3T3 Cells↗

Controlling cell interactions by micropatterning in co-cultures: hepatocytes and 3T3 fibroblasts.

The repair or replacement of damaged tissues using in vitro strategies has focused on manipulation of the cell environment by modulation of cell-extracellular matrix interactions, cell-cell interactions, or soluble stimuli. Many of these environmental influences are easily controlled using macroscopic techniques; however, in co-culture systems with two or more cell types, cell-cell interactions have been difficult to manipulate precisely using similar methods. Although microfabrication has been widely utilized for the spatial control of cells in culture, these methods have never been adapted to the simultaneous co-cultivation of more than one cell type. We have developed a versatile technique for micropatterning of two different cell types based on existing strategies for surface modification with aminosilanes linked to biomolecules and the manipulation of serum content of cell culture media. This co-culture technique allowed manipulation of the initial cellular microenvironment without variation of cell number. Specifically, we were able to control the level of homotypic interaction in cultures of a single cell type and the degree of heterotypic contact in co-cultures over a wide range. This methodology has potential applications in tissue engineering, implant biology, and developmental biology, both in the arena of basic science and optimization of function for technological applications.

3T3 Cells↗

Role of microliths in the aetiology of chronic submandibular sialadenitis: a clinicopathological investigation of 154 cases.

AIMS: Confusion about the aetiology and pathogenesis of chronic submandibular sialadenitis led to the present investigation of 154 cases in which many clinical and histological features were analysed. METHODS AND RESULTS: By far the greatest number of histological factors, namely liths, atrophy, fibrosis, parenchymal inflammation, lymphoid germinal centres, mucous and ciliary metaplasia, salivary extravasation and glycosaminoglycan accumulation, was related to the degree of inflammation, which appears to be of the greatest importance in the aetiology and pathogenesis. Inflammation, atrophy and fibrosis were related to duration of symptoms, which supports the concept of a chronological progression through increasingly severe histological changes. CONCLUSIONS: Inflammation possibly arises from ascending infection in a normal gland and exerts an obstructive and destructive effect on the parenchyma with the development of the related histological changes and a vicious circle involving further ascending infection. Normal glands contain microliths that possibly by localized obstruction cause atrophic foci that are reservoirs for ascending infection. Microliths and liths were unrelated: microliths were related to age as in normal glands whereas liths were related to duration of symptoms and appeared to be secondary to the sialadenitis. Many glands showed minimal changes, which raises the possibility of conservative treatment.

Adolescent↗

Selective adhesion of hepatocytes on patterned surfaces.

Successful development of cell-biased bioartificial liver devices necessitates the establishment of techniques and designs for long-term, stable hepatocellular function and efficient transport of nutrients and wastes within the device. Given the relatively large cell mass that one must consider, one possible solution involves the use of micropatterning technology to sandwich hepatocytes aligned in rows between two micropatterned surfaces. Rows of cells would alternate with hepatocyte-free areas, creating efficient transport channels for fluid flow and nutrient exchange. Ultimately, this type of device could also be used as a three-dimensional construct for investigating a variety of cell-surface, cell-extracellular matrix, and cell-cell interactions. To achieve this goal, one must develop techniques for selectively adhering hepatocytes to solid substrates. In this study, reproducible, selective adhesion of hepatocytes on a glass substrate with large regions of adhesive (AS) and nonadhesive (NAS) surfaces was obtained. The AS had hydrophilic characteristics, enhancing deposition of collagen molecules from an aqueous solution, and subsequent hepatocyte adhesion, whereas the NAS had hydrophobic properties and remained collagen-free and hepatocyte-free. In addition, a reproducible processing technique for obtaining patterns of hepatocytes was developed and optimized, using a surface with a single AS band as a first approximation to a micropatterned device. This was achieved by spincoating an aqueous collagen type I solution (0.1 mg/mL) on a banded surface at 500 rpm for 25 seconds. The morphology and long-term function of the hepatocytes attached to AS in nonbanded and banded surface configurations was assessed by mimicking sandwich culture and was shown to be similar to stable, differentiated sandwich cultures. Mathematical modeling was used to determine critical design criteria for the hypothetical micropatterned device. The oxygen distribution and viscous pressure drop were modeled along a typical microchannel and limited to in vivo values. An optimal channel length of 0.6 cm and a flow rate of 2.0 x 10(-6) mL/s were obtained for a channel of 100 microns in width and 10 microns in height. These values were reasonable in terms of practical implementation.

Albumins↗

Tooth positions in the natural and complete artificial dentitions, with special reference to the incisor teeth: an interactive on-line computer analysis.

A comparison of tooth positions and soft tissue profiles was made between samples of edentulous patients recently provided with new complete dentures, and dentate subjects. The dentures were marked with lead foil on the most prominent incisor teeth and labial surfaces of the flanges. Lateral skull radiographs were made with the jaws in centric relation. The dentate subjects were recorded with the teeth held in centric occlusion. Discrete soft and hard tissue landmarks were identified and recorded using an interactive on-line computer system, enabling linear and angular measurements to be made. A mathematical comparison was produced employing a series of independent t-tests. Composite computer tracings were also produced. Important differences were found between the two groups, in which the influence of the upper lip length, mandibular resorption, and selected overbite were identified as factors determining artificial tooth position.

Adult↗

An interactive computer program for recording and analysing longitudinal cephalometric growth material.

An interactive suite of chained programs is described for recording, measuring, and analysing a large number of longitudinal cephalometric records. The programs are arranged in a logical sequence, and integrated with the hardware, to ensure a smooth and automatic running of the system from data collection to data analysis. The results of the analysis can be presented in suitable numerical and/or graphical forms to show individual as well as mean growth profiles.

Adolescent↗

A comprehensive interactive on-line computer system for research and clinical practice in orthodontics.

An interactive on-line computer system for the measurement of cephalometric radiographs, and facial and plaster cast photographs is described. The details of the hardware (computer and peripherals) and software are provided. Numerical and graphical analysis and presentation of the resultant data are discussed. Many research and clinical applications of the system are outlined.

Cephalometry↗

Nature of relapse after surgical mandibular advancement.

Pre-operative, post-operative and follow-up cephalometric records of 16 cases of mandibular retrognathia treated by surgical mandibular advancement were analysed retrospectively. The results showed good mean stability in the mandibular advancement with variable individual relapse. The pre-operative mandibular plane angle, magnitude of the advancement and post-operative increase in the posterior lower face height were the variables chiefly related to relapse. There appeared to be limited control over the posterior segment which was liable to distraction and rotation. The follow-up changes were time linked. Both mechanical and biological factors are postulated to explain the relapse.

Cephalometry↗

A computer-aided design for orthognathic surgery.

An interactive on-line computer system is described with application in orthognathic surgery. Both the hardware and software of the system are discussed. The application of the system is outlined under the various features of the system's software. The general software collects, stores and analyses graphic data such as from cephalometric radiographs and facial and dental photographs. The specific software handles the planning requirements of orthognathic surgery--diagnosis, treatment planning and prediction of post-operative soft-tissue profile.

Cephalometry↗

A cephalometric analysis of the Le Fort II osteotomy in the adult cleft patient.

A variety of procedures are available for the correction of the secondary cleft deformity. In this study of the use of the Le Fort II osteotomy, hard and soft tissue changes are documented using 40 discrete hard and soft tissue points on each lateral cephalogram. These were compared in the pre-operative, immediate post-operative and follow-up cephalograms. Some of the possible causes for relapse were identified.

Adolescent↗