NRU :: Browsing by Author "Kisaalita, William S."

Browsing by Author "Kisaalita, William S."

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3D Nerve Cell Cultures and Complex Physiological Relevance
(Drug Discovery Today, 2018) Cheng, Xin; Ndyabawe, Kenneth; Asthana, Amish; Kisaalita, William S.
The field of tissue engineering has not yet provided knowledge on which a consensus for the complex physiological relevance (CPR) of neuronal cultures could be established. The CPR of 3D neuronal cultures can have a profound impact on the drug discovery process through the validation of in vitro models for the study of neuropsychiatric and degenerative diseases, as well as screening for neurotoxicity during drug development. Herein, we assemble evidence in support of the potential of [Ca 2+] i oscillation frequency as a CPR outcome that can demonstrate the in vivo-like behavior of 3D cultures and differentiate them from 2D monolayers. We demonstrate that [Ca 2+] i oscillation frequencies in 2D cultures are significantly higher than those found in 3D cultures, and provide a possible molecular explanation.
A Fiber-optic System for Measuring Single Excitation-Dual Emission Fluorescence Ratios in Real Time
(Biotechnology progress, 1992) McCarthy, John F.; Magin, Richard L.; Kisaalita, William S.; Slininger, Patricia J.
The development and subsequent evaluation of a fiber‐optic system for measuring single excitation—dual emission fluorescence ratios in real time is described. The design of the flashlamp excitation source, optics, electronics, and computer software is discussed. The dual emission pH sensitive fluorophore 1, 4‐dihydroxyphthalonitrile (1, 4‐DHPN) was used to test the performance of this system. The flexible design of this modular system permits the use of other single excitation—dual emission fluorophores by simply changing the appropriate optical filters. Upon a single 340–380‐nm excitation, pH‐sensitive emissions were monitored at 488 nm and 434 nm. The ratio of these emissions (488/434) was then computed in real time, for a 2 mM solution of 1, 4‐DHPN, while the pH was titrated over the range 5–9. The nonlinear, system‐dependent, calibration curve of pH versus the ratio of emission wavelengths was empirical fit by a fourth‐order polynomial (r2 = 0.995). Reliable pH measurements in the range 6–8 were obtained using concentrations of 1, 4‐DHPN as low as 50 μM. The standard deviation of pH measurements using a 1 mM solution of 1, 4‐DHPN, near neutral pH, was found to be approximately 0.1 pH unit
A microwell pattern for C17.2 cell aggregate formation with concave cylindrical surface induced cell peeling
(Biomaterials, 2014) Zhang, Li-Guang; Zhong, Dong-Huo; Zhang, Yiguo; Li, Chen-Zhong; Kisaalita, William S.; Wu, Ze-Zhi
We have developed a polydimethylsiloxane (PDMS) pattern with arrays of microwells for the formation of multicellular aggregates by C17.2 neural stem cells. Upon interfacing with the patterns, the neural stem cells would firstly attach to the microwell sidewalls, forming cellular strips on day 1 after plating. For channel connected microwells, cellular strips on the concave semi-cylindrical sidewall surfaces continued among wells and through channels, followed by strip peeling due to prestress arising from actin filaments and assembly of suspending cellular aggregates within the microwells in the following 1–2 days. Our results also suggested that a small microwell diameter of 80 and 100 μm and a narrow channel width of 20 μm would facilitate the aggregate formation among the structural dimensions tested. Finite element method (FEM) simulation revealed that cellular strips on the semi-cylindrical sidewall surfaces peeled under significantly smaller prestresses (critical peeling prestress, CPP), than cells on flat substrates. However, the CPP by itself failed to fully account for the difference in aggregate inducing capability among the patterns addressed, suggesting cell growth behaviors might play a role. This study thus justified the current patterning method as a unique and practical approach for establishing 3D neural stem cell-based assay platform
A single magnetic field exposure system for sequential investigation of real time and downstream cellular responses
(Bioelectromagnetics, 2004) Rao, Raj R.; Kisaalita, William S.
To be able to correlate real time membrane potential or ion flux changes with further downstream gene transcription responses due to extremely low frequency (ELF) electromagnetic field (EMF) exposure, we devised an experimental system consisting of a pair of symmetric circular coils. This system can be used on an inverted microscope stage (real time signaling) as well as inside controlled environment incubators (gene transcription end points). The system includes a unique, custom made switch box for blinding the experimental staff and a power amplifier. We report herein the design and characterization of the system with respect to parameters considered important in in vitro ELF–EMF exposure studies, including linear magnetic field distribution, compensation for microscope objective lens interference, heating effects of the coils, and harmonic content of the signals. Bioelectromagnetics 25:27–32, 2004. © 2003 Wiley-Liss, Inc.
Administration of BDNF/ginsenosides combination enhanced synaptic development in human neural stem cell
(Journal of neuroscience methods, 2011) Wang, Lina; Kisaalita, William S.
Ginsenosides Rg1 and Rb1, major pharmacologically active ingredients from Ginseng, the root of Panax ginseng C.A. Meyer (Araliaceae), were applied in the differentiation media for human neural stem cells (hNSCs), together with brain-derived neurotrophic factor (BDNF), a commonly used compound for neural stem cell (NSC) differentiation. Cell locomotion and neurite extension were observed by time-lapse microscopy and analyzed by ImageJ software. The expression of synaptic formation was confirmed by immunostaining of synaptophysin (SYN) or/and the co-localization of synapsin I and microtubule associated protein-2 (MAP-2). Effects of cell density on neural differentiation were also examined. Results have shown that administration of BDNF/ginsenosides (Rg1 and Rb1) combination in differentiation medium promoted cell survival, enhanced neurite outgrowth and synaptic marker expression during differentiation. High cell density enhanced synaptic marker expression in BDNF/ginsenosides combination medium. In all, this study established a condition for hNSCs synaptic development in early differentiation, which is a crucial step in applying this cell line in neural network-based assay.
Anthropometric characteristics of female smallholder farmers of Uganda – Toward design of labor-saving tools
(Applied Ergonomics, 2016) Mugisa, Dana J.; Katimbo, Abia; Sempiira, John E.; Kisaalita, William S.
Sub-Saharan African women on small-acreage farms carry a disproportionately higher labor burden, which is one of the main reasons they are unable to produce for both home and the market and realize higher incomes. Labor-saving interventions such as hand-tools are needed to save time and/or increase productivity in, for example, land preparation for crop and animal agriculture, post-harvest processing, and meeting daily energy and water needs. Development of such tools requires comprehensive and content-specific anthropometric data or body dimensions and existing databases based on Western women may be less relevant. We conducted measurements on 89 women to provide preliminary results toward answering two questions. First, how well existing databases are applicable in the design of hand-tools for sub-Saharan African women. Second, how universal body dimension predictive models are among ethnic groups. Our results show that, body dimensions between Bantu and Nilotic ethnolinguistic groups are different and both are different from American women. These results strongly support the need for establishing anthropometric databases for sub-Saharan African women, toward hand-tool design.
Assessment of Murine Neuroblastoma (N1E-115) Resting Membrane Potential by Confocal Microscopy
(Journal of Fluorescence, 1996) Hernandez, Miguel; Kisaalita, William S.; Farmer, Mark A.
Digital imaging (confocal microscopy) and a slow potentiometric dye (tetramethylrhodamine methyl ester) were used to assess the resting membrane potential (V m) of murine neuroblastoma cells (N1E-115). The averageV m was found to be −64.0±2.0 mV. The difference between this and the previously reported higher values was attributed to the use of glass microelectrode techniques that probably caused mechanical injury to the cell membranes: Digital imaging of N1E-115V m was found to be sensitive, reproducible, fast, and simple.
Biogas-powered evaporative cooling for smallholder dairy farmers’ evening milk: Zeolite characterization and regeneration
(Sustainable Energy Technologies and Assessments, 2019) Ndyabawe, Kenneth; Brush, Ryan; Ssonko, Richard E.; Kisaalita, William S.
A low-capacity (15.5-L) evaporative cooler, utilizing zeolite as an adsorbent, for saving smallholder dairy farmers’ evening milk, was not well received due several factors, addressed in the second generation higher capacity (>50 L) design. Like the first generation the new design uses zeolite and is powered by biogas. The first development step was to establish a protocol for testing commercially available zeolites to determine zeolite suitability for use in the design and to characterize the performance of a zeolite biogas-powered regenerator. Using an in-house developed protocol, zeolite beads of diameters 2.5–5 mm were tested and we found that 10 angstrom (Å) zeolites have a higher water adsorption capacity (6–7%) in comparison to 3 Å zeolite beads, suggesting that larger pore zeolites provide higher cooling capacity. However, large pore zeolite beads showed up to 18 times variance in repeat water adsorption compared to 3 Å zeolite, indicating that zeolite of small pores may provide more cooling cycles. Our biogas powered regenerator achieved and maintained the regeneration temperature of 200 °C when tested with both propane and biogas. This result affirms that our system can be operated on the farm using biogas as the energy source.
Biomarkers for simplifying HTS 3D cell culture platforms for drug discovery: the case for cytokines
(Drug discovery today, 2011) Lai, Yinzhi; Asthana, Amish; Kisaalita, William S.
In this review, we discuss the microenvironmental cues that modulate the status of cells to yield physiologically more relevant three-dimensional (3D) cell-based high throughput drug screening (HTS) platforms for drug discovery. Evidence is provided to support the view that simplifying 3D cell culture platforms for HTS applications calls for identifying and validating ubiquitous three-dimensionality biomarkers. Published results from avascular tumorigenesis and early stages of inflammatory wound healing, where cells transition from a two-dimensional (2D) to 3D microenvironment, conclusively report regulation by cytokines, providing the physiological basis for focusing on cytokines as potential three-dimensionality biomarkers. We discuss additional support for cytokines that comes from numerous 2D and 3D comparative transcriptomic and proteomic studies, which generally report upregulation of cytokines in 3D compared with 2D culture counterparts.
Biophysical microenvironment and 3D culture physiological relevance
(Drug discovery today, 2013) Asthana, Amish; Kisaalita, William S.
Force and substrate physical property (pliability) is one of three well established microenvironmental factors (MEFs) that may contribute to the formation of physiologically more relevant constructs (or not) for cell-based high-throughput screening (HTS) in preclinical drug discovery. In 3D cultures, studies of the physiological relevance dependence on material pliability are inconclusive, raising questions regarding the need to design platforms with materials whose pliability lies within the physiological range. To provide more insight into this question, we examine the factors that may underlie the studies inconclusiveness and suggest the elimination of redundant physical cues, where applicable, to better control other MEFs, make it easier to incorporate 3D cultures into state of the art HTS instrumentation, and reduce screening costs per compound.
Biosensor Standards Requirements
(Biosensors & Bioelectronics, 1992) Kisaalita, William S.
Since biosensor applications are springing up in many places, it should be the wish of all biosensor manufacturers, their customers, as well as those in academia, that the same language and requirements are used to describe similar products. In order to find answers to the question as to whether the time is right to consider developing standards for biosensors, and if so, what the priority areas might be, we have conducted a survey of biosensor users and/or manufacturers. It is the purpose of this article to report our survey results.
Brain-on-a-Chip Device for Modeling Multiregional Networks
(ACS Biomaterials Science & Engineering, 2020) Ndyabawe, Kenneth; Cipriano, Michael; Zhao, Wujun; Haidekker, Mark; Yao, Kun; Mao, Leidong; Kisaalita, William S.
Animal models are frequently used in drug discovery because they represent a mammalian in vivo model system, they are the closest approximation to the human brain, and experimentation in humans is not ethical. Working with postmortem human brain samples is challenging and developing human in vitro systems, which mimic the in vivo human brain, has been challenging. However, the use of animal models in drug discovery for human neurological diseases is currently under scrutiny because data from animal models has come with variations due to genetic differences. Evidence from the literature suggests that techniques to reconstruct multiple neurotransmission projections, which characterize neurological disease circuits in humans, in vitro, have not been demonstrated. This paper presents a multicompartment microdevice for patterning neurospheres and specification of neural stem cell fate toward networks of multiple neuronal phenotypes. We validated our design by specification of human neural stem cells to dopaminergic and GABAergic neurons in different compartments of the device, simultaneously. The neurospheres formed unrestricted robust neuronal circuits between arrays of neurospheres in all compartments of the device. Such a device design may provide a basis for formation of multineurotransmission circuits to model functional connectivity between specific human brain regions, in vitro, using human-derived neural stem cells. This work finds relevance in neurological disease modeling and drug screening using human cell-based assays and may provide the impetus for shifting from animal-based models.
Capstone Design Experiences Across National and Cultural Borders: Course Development
(American Society for Engineering Education, 2002) Kisaalita, William S.
It is well recognized that the world is becoming a global village. Today's undergraduates will occupy workplaces and communities that have been transformed by globalization; they must learn to make connections across disciplinary, national, and cultural borders. To address the challenge of providing exposure to global/international dimensions without sacrificing technical content, we have previously proposed to use case studies in sophomore engineering science classes. In this paper, we extend the concept to capstone engineering design courses by focusing on solutions to global/international problems. Additionally, a Web-based instructional tool (WebCT) is used to enhance global/international awareness as well as intercultural communication skills. WebCT is also used to “take students there,” through movie clip
Cell adhesion and locomotion on microwell-structured glass substratesCell adhesion and locomotion on microwell-structured glass substrates :
(Biointerfaces, 2011) Guan, Yanbin; Kisaalita, William S.
The purpose of this study was to investigate the effect of microstructured material surface on cell adhesion and locomotion in real-time. ArF excimer laser direct-writing ablation was used to fabricate microwell patterns with precise control of size and spacing on glass. The influence of the ablation process parameters (laser fluence, pulse number and repetition rate) on the micromachining quality (depth, width, aspect ratio and edge effects) of the microwells was established. Human fibroblast cells, as an example of anchorage-dependent cells, were seeded onto the microstructured glass substrate and time-lapse microscopy was used to study cell adhesion and locomotion. The interaction with microstructured materials resulted in fibroblast cell repulsion and the cells exhibited a higher locomotion speed (75.77±3.36μm/h) on the structures in comparison with plane glass control (54.01±15.53μm/h). Further studies are needed to firmly establish the potential of microstructuring, for example, in elongating the life spans of implantable devices.
Characterization of 3-D collagen hydrogels for functional cell-based biosensing
(Biosensors and Bioelectronics, 2004) Mao, Chen; Kisaalita, William S.
To address the growing demand for functional cell-based assay technologies with accelerated drug discovery applications, we have proposed the use of human neuroblastoma cells (IMR-32) immobilized in three-dimensional (3-D) collagen hydrogel matrices. The gel protects weakly adherent cells from fluid mechanical forces while providing a more physiologically relevant 3-D environment. Hydrogels made up of collagen, between 0.5 and 1.0mg/ml, exhibited mechanical stability adequate to withstand fluid mechanical forces (<0.11mN) typical of automated commercial fluid transfer equipment. Collagen-entrapped cells visualized with the aid of confocal microscopy and a potentiometric-sensitive dye, TMRM, exhibited round morphology in comparison to flat morphology typical of cells in two-dimensional (2-D) monolayer cultures. Morphological differentiation characterized by neurite extension and cell aggregation was observed for both 2-D and 3-D cultures. Differentiated IMR-32 cells failed to develop a resting membrane potential typical of excitable cells. Free intracellular calcium was monitored with Calcium Green-1. Depolarization-induced Ca2+ influx was only observed with differentiated 3-D cells unlike 2-D cells, where calcium flux was observed in both differentiated and undifferentiated cells. Taken together, the results revealed that collagen hydrogels (0.5mg/ml collagen) were suitable structural supports for weakly adherent cells. However, for voltage-dependent calcium channel function applications, further investigations are needed to explain the difference between 2-D monolayer and 3-D collagen-entrapped cells
Characterization of micropatterned nanofibrous scaffolds for neural network activity readout for high-throughput screening
(Applied Biomaterials, 2010) Wang, Lina; Kisaalita, William S.
Micropatterns were fabricated in nanofibrous poly‐L‐lactic acid (PLLA) films by laser micromachining and the resulting scaffolds were characterized with respect to architecture, thermal, mechanical, and mass transport properties. Also, human neural stem cells were successfully cultured in these micropatterned nanofibrous scaffolds (MNFSs). The scaffolds were incorporated in high‐density well plates (e.g., 96‐well plates), creating a platform for high‐throughput screening of drugs with physiologically more relevant networked neural cultures. Through mathematical modeling of the transport of model stimulants, the feasibility of stimulating neural networks cultured in MNFSs was demonstrated. More work is needed to establish biological network activity–MNFS architecture relationships. © 2010 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 2010.
Combining thermization and evaporative cooling toward milk freshness preservation at the smallholder farm level
(Journal of Food Process Engineering, 2020) Sempiira, Edison J.; Mugisa, Dana J.; Galiwango, Joseph; Kisaalita, William S.
Thermization and low‐cost evaporative cooling have been seamlessly combined (evakuuling process) to preserve milk freshness for next day market. For proper regulation, a more exact profile of thermized milk in the hands of smallholders is needed. The purpose of this paper was to establish the milk quality‐profile of “evakuuled” milk. Thermization was done using temperatures between 55 and 70°C. Evakuuled milk quality‐profile (total viable and psychotrophic microbial counts, fat, protein, and pH) was compared to chilled and fresh milk. A consumer sensory test was also performed to evaluate consumer preference. The results showed that the quality‐profile of milk thermized at 65°C, followed by evaporative cooling (evakuuled) was indistinguishable from chilled and fresh milk. A consumer preference test scored evakuuled, fresh and chilled milks on average equal in terms of taste, appearance, aroma, and general acceptability. Practical applications The finding of evakuuled milk having the same quality attributes as fresh or chilled milk and meeting consumer preference in terms of aroma, taste appearance and general acceptability has potential practical significance. These results provide evidence in support of inclusion of evakuuled milk in the East African raw milk standard.
Comparative evaluation of the susceptibility of neuronal (N1E-115) and non-neuronal (HeLa) cells to acetylsalicylic acid (ASA) cytotoxicity by confocal microscopy
(Toxicology in vitro, 1996) Hernandez, M.; Kisaalita, William S.
A voltage-sensitive probe, tetramethylrhodamine methyl ester (TMRM) and digital imaging (confocal microscopy) were used to quantify differential membrane potential alteration in neuronal (murine neuroblastoma, N1E-115) and non-neuronal (human epithelial-like, HeLa) cells, after 1 and 24 hr of exposure to a knownin vivo neurotoxic agent, acetylsalicylic acid (ASA), as a first step in substantiating the relevance of alteration in resting membrane potential (Vm) as a neurotoxic endpoint. After 1 hr of exposure, ASA (5.0 mm) hyperpolarized both HeLa and N1E-115 cells. Vm decreased from −57.6mV± 2.8 (n= 20cells)to−74.7mV± 1.9 (n= 20cells) and from −64.0mV± 2.1 (n= 20cells)to−82.5mV± 3.4 (n= 20cells) for HeLa and N1E-115 cells, respectively. The extent of hyperpolarization was found to be ASA concentration dependent. There was no significant difference (P < 0.05) between the cell lines with respect to ASA sensitivity, suggesting that under these experimental conditions, ASA exhibited no selective cytotoxic activity for the neuronal cells. In comparison with control cultures, 24-hr ASA (5.0 mm) exposure did not affect the surviving cell Vm. The results of the present study were inconclusive with respect to the suitability of Vm alteration as an indicator of neurotoxic potential.
Comparative Feasibility Analysis of Alternative Renewable Energy Sources for Small Milk Cooling Plants of Southwestern Uganda
(Africa & Latin America, 2006) Kisaalita, William S.; Franklin, Justin; Tippie, Allyson; Boyer, Brad; Faircloth, William
The dairy industry in Uganda faces numerous challenges, one of which is the high cost associated with fossil fuel-driven milk cooling processes at locations without grid electricity. In this paper, alternative renewable energy sources, including solar and biogas, are compared to the existing diesel generator power source for a 3,000 L-capacity plant. Based on several economic indicators, the solar (evacuated tube) source coupled to an absorption refrigeration unit was found to be the most economical alternative for southwestern locations
Cultural Influences in Women-Friendly Labor- Saving Hand Tool Designs :
(Human Factors, 2016) Kisaalita, William S.; Katimbo, Abia; Sempiira, Edison J.; Mugisa, Dana J.
One of the reasons behind the gender asset gap among Sub-Saharan African women is the higher labor burden these women face, making it difficult for them to produce for the home and markets. Hand tools are the simplest form and therefore the best first step to address this problem. But designing women-friendly (sustainable) hand tools calls for better understanding of the low-resource settings where these women reside.

Browsing by Author "Kisaalita, William S."

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