The Indonesian archipelago comprises more than 17,000 islands, inhabited by ~200 million people constituting more than 350 recognizable ethnic and tribal groups which can be classified into two broad ethno-linguistic groups [the Austronesian (AN) and non-Austronesian (NAN) speaking peoples] and 3 physical anthropology groups (Deutero Malay, Proto Malay and Papuan). The origins of these groups are of considerable anthropological interest. The anthropology of Indonesia is extremely complex and still controversial. The present populations of Indonesia show very great diversity. The data presented below result from an investigation of the Fragile X A syndrome and the size and distribution of alleles at fragile sites on the X chromosome among Javanese males with developmental disability (DD) and unselected males from 10 major Indonesian ethnic groups. The Fragile X syndrome is caused by expansion of a CGG trinucleotide repeat array in the 5' untranslated region of the FMR-1 gene at Xq27.3. Normal X chromosomes have between 6-54 CGG trinucleotide repeats, whereas premutation alleles have 55-230 and full mutation alleles more than 230 repeats. In a study of predominantly Caucasian males with intellectual disability, the prevalence of Fragile X syndrome is estimated to be approximately 1:4,000. FRAXE mental retardation syndrome is caused by an expansion of a GCC trinucleotide repeat in the 5'UTR of FMR2 gene located 600 kb telomeric to FMR1. The prevalence of FMR2 is 1-2 per 100,000 live births. FMR2 common alleles consist of 11-30 GGC repeats; intermediate alleles between 31-60 GCC repeats; premutation alleles with 61-200 repeats and full mutation alleles have over 200 repeats with attendant methylation of the repeat array The first Indonesian screening program aimed at determining the presence and prevalence of fragile XA syndrome among individuals with mild DD (IQ above 50) from special schools (N=205) and isolated areas (N=50) of Java was undertaken in 1994-1996 by cytogenetic and molecular studies. In this first study 4 fragile X positive children were found among 255 males with DD. The estimated prevalence of fragile-X in males with mild DD from special schools was 1.95% (5/205) and the overall prevalence was 1.57% (4/255). The number of trinucleotide repeats in the 5' untranslated regions of the FMR1 and FMR2 genes were determined by PCR in 254 Fragile XA-negative Javanese male children with DD. The distribution of FMR1 and FMR2 trinucleotide repeat alleles was found to be significantly different in the Indonesian population with DD compared to that in equivalent Caucasian populations. The trimodal distribution of Indonesian FMR1 alleles (29, 30 and 36 repeats) is largely in agreement with findings from other Asian populations). This provides supportive evidence that the origin of Indonesians could be the same as that of the Chinese and Japanese. Sequence analysis was performed on the trinucleotide repeat arrays of the 27 individuals' FMR1 alleles in the 'grey zone' (35-52 repeats). The identification of 16 unrelated individuals with a (CGG)36 allele that also contains a (CGG)6 segment [(CGG)9AGG(CGG)9AGG(CGG)6 AGG(CGG)9 or 9A9A6A9 pattern] is in agreement with earlier observations in the Japanese population. It is proposed that this FMR1 array pattern may be specific for Asian populations and that Javanese and Japanese populations may have arisen from a single progenitor population. The presence of pure 25, 33 and 34 CGGs in FMR1 alleles with 36, 44 and 45 repeats respectively, suggests that these may represent alleles at high risk for instability and may therefore be at early stages of expansion to a premutation. The lack of the characteristic (CGG)6 in all three alleles with ?? 25 pure CGG arrays suggests that the most common Asian 36 repeat allele is not predisposed to slippage expansion. Seven of the 8 alleles with 36 CGG repeats could be sequenced. Seven of 36 CGG repeats FMR1 alleles from the Hiri population has been sequenced and 4 alleles indicated 9A9A6A9 pattern, 1 sample with 10A25 pattern Two of the remaining alleles showed 12A6A6A9 structure, which consisted of a tandem duplication of the (CGG)6 segment. The presence of a tandem duplication of (CGG)6 segments has never been reported in any other population. The other major findings of this study are that FRAXE syndrome is a rare cause of developmental disability in this predominantly-Javanese population. The most common FMR2 (GCC)20 allele in this selected Asian population is significantly longer than that previously reported for Caucasian populations. There was a weak correlation between the overall length of the FMR1 and FMR2 repeat arrays within the normal range (Spearman's Rank Correlation = 0.130, p-value=0.042) in the Indonesian population, which have been no previous associations reported for alleles within the normal range. One approach to studying the origins of the human populations is to study the genetic structure of polymorphic alleles such as those at the FMR1 locus and its linked microsatellite markers DXS548 and FRAXAC1. Length polymorphisms of the FMR1 gene (CGG)n repeat array, DXS548 and FRAXAC1 were studied in a total of 1,008 unselected males from 10 different Indonesian ethnic groups. FMR1 alleles were identified ranging from 8 to 57 CGG repeats. The most common CGG repeat allele was 29 (45.6%) followed by 30 (27.4%) and 36 repeats (8.0%). One hundred and forty four grey zone (3-52 CGG) alleles were found in the study population. Four people of the same ethnic group from an isolated island in Eastern Indonesia (Hiri, Ternate), a representative of the NAN ethnolinguistic group, had CGG repeat lengths of 55-57. The prevalence of these alleles is estimated to be 3.3% (4/120) in the population of Hiri or 0.4% (4/1008) of whole Indonesian population. Thirteen different alleles were found at the DXS548 locus, of which allele numbers 7 [194 bp] (44.1%), 6.5 [195bp] (43.5%) and 6 [196bp] (7.5%) are the most common. Seven rare alleles, some of which have not been previously found in Asian peoples were also identified (190, 191,192, 193, 197,198, 199, 202, 204 and 206) and accounted for 3.9% of the total. The odd number alleles were dominantly found in this study whereas almost none found in Caucasian. The finding of many "odd numbered" alleles DXS548 has never been found in other Asian population and has only been documented extremely rarely in Caucasians and Africans. Five different alleles of FRAXAC1 identified with alleles D [106 bp] (62.2%) and C [108bp] (35.6%) accounting for 97.8% of FRAXAC1 alleles in the population. Three rare alleles (104, 110, 112 bp = 2.2%) were identified that have not been previously found in other Asian populations (1-3). There is a striking linkage disequilibrium of FMR1 alleles with FRAXAC1 (p=0.0001), 88% of 29 (CGG)n repeats alleles associated with FRAXAC1 allele D (106bp) versus only 17% with the 30 (CGG)n repeat alleles, which is in agreement with other studies. The value of D' was calculated to be 0.7. The longer alleles of both DXS548 and FRAXAC1 were found mostly in the NAN ethnolinguistic group. Moreover the Irian Jaya people also showed a higher percentage of people with 30 CGG repeats and the 108 bp FRAXAC1. The Eastern Indonesian NAN groups demonstrate a different genetic background probably due to the contribution of Melanesian peoples. The Analysis of Molecular Variance (AMOVA) identified that the vast majority of genetic diversity occurs within, rather than between, ethnic groups. These data are consistent with a model where there is sufficient migration (~20 per generation) between populations to minimise differentiation of population through genetic drift. The results obtained are consistent with three clusters of populations that share similar allele frequencies at the fragile X locus. The most clearly defined cluster is based in the east of Indonesia and includes the two Irian populations, Minahasans and Hiri. A surprising finding was that the Minahasan who are Deutero-Malay in origin and physical appearance are genetically closer to the Irianese. This may reflect the admixture of Melanesian alleles or other eastern Indonesian alleles as a result of their geographic location in that part of Indonesia. The second major cluster is largely based in the west of the country and is composed of the following Deutero-Malay populations; Javanese, Balinese, Acehnese but which also includes people from Ternate (not including those from Hiri). Using Delta Mu and Nei's genetic distance for FMR1 locus in this study the Javanese were shown to have the closest distance to Balinese which is consistent with anthropological data and with published data. The third group is a "western and central" group composed of Bimanese, Dayak and Sundanese who share some features of the western and eastern clusters but mostly resemble the western Indonesian populations. Bima is located in the lesser Sunda in between west Indonesia and east Indonesia. The Bimanese are of mixed Deutero & Proto Malay origin that is consistent with their geographic location. The Bataks are distinctive and sit somewhat apart in this scheme. In this study, Bataks were found not to resemble the other Proto-Malay group studied (the Dayak). The Dayaks were found to have fewer alleles than the Bataks at FRAXAC1 and DXS548. In all four methods of calculating genetic distance Bataks showed a large genetic distance to almost all other ethnic groups. There are differences in allele frequency between east and west Indonesia as well as other Asian nations, but the genetic similarities between these groups are also very impressive. The findings from this study are consistent with other genetic anthropological evidence that the people of Indonesia have the same origin as North-east Asian groups. This model is referred to as the "express train from Taiwan" in which the Austronesian speakers are proposed to have radiated from Taiwan bringing the Malayo-Polynesian language group to the Philippines, Borneo and Sulawesi around 5000-4500 B.P.E. However Richards et al.(1998) have used the diversity in the mtDNA D Loop to propose an alternative to the "express train" model. The "two train7quot; model proposes that the Austronesian languages originated within eastern Indonesia during the Pleistocene era and spread through Melanesia and into the remote Pacific within the past 6,000 years. Unfortunately the high migration rates between population groups that were demonstrated in this thesis and the known migration patterns of populations through Indonesia preclude determining whether the observed allelic heterogeneity is a function of the original population or due to the admixture of several gene pools in more recent times.

Erlangen, Nürnberg, Universiẗat, Diss., 2007. / Enth. 1 Sonderabdr. aus: Journal of investigative dermatology ; 124. 2005. - Beitr. teilw. dt., teilw. engl.

The Indonesian archipelago comprises more than 17,000 islands, inhabited by ~200 million people constituting more than 350 recognizable ethnic and tribal groups which can be classified into two broad ethno-linguistic groups [the Austronesian (AN) and non-Austronesian (NAN) speaking peoples] and 3 physical anthropology groups (Deutero Malay, Proto Malay and Papuan). The origins of these groups are of considerable anthropological interest. The anthropology of Indonesia is extremely complex and still controversial. The present populations of Indonesia show very great diversity. The data presented below result from an investigation of the Fragile X A syndrome and the size and distribution of alleles at fragile sites on the X chromosome among Javanese males with developmental disability (DD) and unselected males from 10 major Indonesian ethnic groups. The Fragile X syndrome is caused by expansion of a CGG trinucleotide repeat array in the 5' untranslated region of the FMR-1 gene at Xq27.3. Normal X chromosomes have between 6-54 CGG trinucleotide repeats, whereas premutation alleles have 55-230 and full mutation alleles more than 230 repeats. In a study of predominantly Caucasian males with intellectual disability, the prevalence of Fragile X syndrome is estimated to be approximately 1:4,000. FRAXE mental retardation syndrome is caused by an expansion of a GCC trinucleotide repeat in the 5'UTR of FMR2 gene located 600 kb telomeric to FMR1. The prevalence of FMR2 is 1-2 per 100,000 live births. FMR2 common alleles consist of 11-30 GGC repeats; intermediate alleles between 31-60 GCC repeats; premutation alleles with 61-200 repeats and full mutation alleles have over 200 repeats with attendant methylation of the repeat array The first Indonesian screening program aimed at determining the presence and prevalence of fragile XA syndrome among individuals with mild DD (IQ above 50) from special schools (N=205) and isolated areas (N=50) of Java was undertaken in 1994-1996 by cytogenetic and molecular studies. In this first study 4 fragile X positive children were found among 255 males with DD. The estimated prevalence of fragile-X in males with mild DD from special schools was 1.95% (5/205) and the overall prevalence was 1.57% (4/255). The number of trinucleotide repeats in the 5' untranslated regions of the FMR1 and FMR2 genes were determined by PCR in 254 Fragile XA-negative Javanese male children with DD. The distribution of FMR1 and FMR2 trinucleotide repeat alleles was found to be significantly different in the Indonesian population with DD compared to that in equivalent Caucasian populations. The trimodal distribution of Indonesian FMR1 alleles (29, 30 and 36 repeats) is largely in agreement with findings from other Asian populations). This provides supportive evidence that the origin of Indonesians could be the same as that of the Chinese and Japanese. Sequence analysis was performed on the trinucleotide repeat arrays of the 27 individuals' FMR1 alleles in the 'grey zone' (35-52 repeats). The identification of 16 unrelated individuals with a (CGG)36 allele that also contains a (CGG)6 segment [(CGG)9AGG(CGG)9AGG(CGG)6 AGG(CGG)9 or 9A9A6A9 pattern] is in agreement with earlier observations in the Japanese population. It is proposed that this FMR1 array pattern may be specific for Asian populations and that Javanese and Japanese populations may have arisen from a single progenitor population. The presence of pure 25, 33 and 34 CGGs in FMR1 alleles with 36, 44 and 45 repeats respectively, suggests that these may represent alleles at high risk for instability and may therefore be at early stages of expansion to a premutation. The lack of the characteristic (CGG)6 in all three alleles with ?? 25 pure CGG arrays suggests that the most common Asian 36 repeat allele is not predisposed to slippage expansion. Seven of the 8 alleles with 36 CGG repeats could be sequenced. Seven of 36 CGG repeats FMR1 alleles from the Hiri population has been sequenced and 4 alleles indicated 9A9A6A9 pattern, 1 sample with 10A25 pattern Two of the remaining alleles showed 12A6A6A9 structure, which consisted of a tandem duplication of the (CGG)6 segment. The presence of a tandem duplication of (CGG)6 segments has never been reported in any other population. The other major findings of this study are that FRAXE syndrome is a rare cause of developmental disability in this predominantly-Javanese population. The most common FMR2 (GCC)20 allele in this selected Asian population is significantly longer than that previously reported for Caucasian populations. There was a weak correlation between the overall length of the FMR1 and FMR2 repeat arrays within the normal range (Spearman's Rank Correlation = 0.130, p-value=0.042) in the Indonesian population, which have been no previous associations reported for alleles within the normal range. One approach to studying the origins of the human populations is to study the genetic structure of polymorphic alleles such as those at the FMR1 locus and its linked microsatellite markers DXS548 and FRAXAC1. Length polymorphisms of the FMR1 gene (CGG)n repeat array, DXS548 and FRAXAC1 were studied in a total of 1,008 unselected males from 10 different Indonesian ethnic groups. FMR1 alleles were identified ranging from 8 to 57 CGG repeats. The most common CGG repeat allele was 29 (45.6%) followed by 30 (27.4%) and 36 repeats (8.0%). One hundred and forty four grey zone (3-52 CGG) alleles were found in the study population. Four people of the same ethnic group from an isolated island in Eastern Indonesia (Hiri, Ternate), a representative of the NAN ethnolinguistic group, had CGG repeat lengths of 55-57. The prevalence of these alleles is estimated to be 3.3% (4/120) in the population of Hiri or 0.4% (4/1008) of whole Indonesian population. Thirteen different alleles were found at the DXS548 locus, of which allele numbers 7 [194 bp] (44.1%), 6.5 [195bp] (43.5%) and 6 [196bp] (7.5%) are the most common. Seven rare alleles, some of which have not been previously found in Asian peoples were also identified (190, 191,192, 193, 197,198, 199, 202, 204 and 206) and accounted for 3.9% of the total. The odd number alleles were dominantly found in this study whereas almost none found in Caucasian. The finding of many "odd numbered" alleles DXS548 has never been found in other Asian population and has only been documented extremely rarely in Caucasians and Africans. Five different alleles of FRAXAC1 identified with alleles D [106 bp] (62.2%) and C [108bp] (35.6%) accounting for 97.8% of FRAXAC1 alleles in the population. Three rare alleles (104, 110, 112 bp = 2.2%) were identified that have not been previously found in other Asian populations (1-3). There is a striking linkage disequilibrium of FMR1 alleles with FRAXAC1 (p=0.0001), 88% of 29 (CGG)n repeats alleles associated with FRAXAC1 allele D (106bp) versus only 17% with the 30 (CGG)n repeat alleles, which is in agreement with other studies. The value of D' was calculated to be 0.7. The longer alleles of both DXS548 and FRAXAC1 were found mostly in the NAN ethnolinguistic group. Moreover the Irian Jaya people also showed a higher percentage of people with 30 CGG repeats and the 108 bp FRAXAC1. The Eastern Indonesian NAN groups demonstrate a different genetic background probably due to the contribution of Melanesian peoples. The Analysis of Molecular Variance (AMOVA) identified that the vast majority of genetic diversity occurs within, rather than between, ethnic groups. These data are consistent with a model where there is sufficient migration (~20 per generation) between populations to minimise differentiation of population through genetic drift. The results obtained are consistent with three clusters of populations that share similar allele frequencies at the fragile X locus. The most clearly defined cluster is based in the east of Indonesia and includes the two Irian populations, Minahasans and Hiri. A surprising finding was that the Minahasan who are Deutero-Malay in origin and physical appearance are genetically closer to the Irianese. This may reflect the admixture of Melanesian alleles or other eastern Indonesian alleles as a result of their geographic location in that part of Indonesia. The second major cluster is largely based in the west of the country and is composed of the following Deutero-Malay populations; Javanese, Balinese, Acehnese but which also includes people from Ternate (not including those from Hiri). Using Delta Mu and Nei's genetic distance for FMR1 locus in this study the Javanese were shown to have the closest distance to Balinese which is consistent with anthropological data and with published data. The third group is a "western and central" group composed of Bimanese, Dayak and Sundanese who share some features of the western and eastern clusters but mostly resemble the western Indonesian populations. Bima is located in the lesser Sunda in between west Indonesia and east Indonesia. The Bimanese are of mixed Deutero & Proto Malay origin that is consistent with their geographic location. The Bataks are distinctive and sit somewhat apart in this scheme. In this study, Bataks were found not to resemble the other Proto-Malay group studied (the Dayak). The Dayaks were found to have fewer alleles than the Bataks at FRAXAC1 and DXS548. In all four methods of calculating genetic distance Bataks showed a large genetic distance to almost all other ethnic groups. There are differences in allele frequency between east and west Indonesia as well as other Asian nations, but the genetic similarities between these groups are also very impressive. The findings from this study are consistent with other genetic anthropological evidence that the people of Indonesia have the same origin as North-east Asian groups. This model is referred to as the "express train from Taiwan" in which the Austronesian speakers are proposed to have radiated from Taiwan bringing the Malayo-Polynesian language group to the Philippines, Borneo and Sulawesi around 5000-4500 B.P.E. However Richards et al.(1998) have used the diversity in the mtDNA D Loop to propose an alternative to the "express train" model. The "two train7quot; model proposes that the Austronesian languages originated within eastern Indonesia during the Pleistocene era and spread through Melanesia and into the remote Pacific within the past 6,000 years. Unfortunately the high migration rates between population groups that were demonstrated in this thesis and the known migration patterns of populations through Indonesia preclude determining whether the observed allelic heterogeneity is a function of the original population or due to the admixture of several gene pools in more recent times.

Thesis (Ph. D.)--Ohio State University, 2006. / Title from first page of PDF file. Includes bibliographical references (p. 178-197).

The interplay between learning and technology is a growing field that is often referred to as Technology Enhanced Learning (TEL). Within this context, learning applications are software components that are useful for learning purposes, such as textbook replacements, information gathering tools, communication and collaboration tools, knowledge modeling tools, rich lab environments that allows experiments etc. When developing learning applications, the choice of technology depends on many factors. For instance, who and how many the intended end-users are, if there are requirements to support in-application collaboration, platform restrictions, the expertise of the developers, requirements to inter-operate with other systems or applications etc. This thesis provides guidance on a how to develop learning applications based on Semantic Web technology. The focus on Semantic Web technology is due to its basic design that allows expression of knowledge at the web scale. It also allows keeping track of who said what, providing subjective expressions in parallel with more authoritative knowledge sources. The intended readers of this thesis include practitioners such as software architects and developers as well as researchers in TEL and other related fields. The empirical part of the this thesis is the experience from the design and development of two learning applications and two supporting frameworks. The first learning application is the web application Confolio/EntryScape which allows users to collect files and online material into personal and shared portfolios. The second learning application is the desktop application Conzilla, which provides a way to create and navigate a landscape of interconnected concepts. Based upon the experience of design and development as well as on more theoretical considerations outlined in this thesis, three major obstacles have been identified: The first obstacle is: lack of non-expert and user friendly solutions for presenting and editing Semantic Web data that is not hard-coded to use a specific vocabulary. The thesis presents five categories of tools that support editing and presentation of RDF. The thesis also discusses a concrete software solution together with a list of the most important features that have crystallized during six major iterations of development. The second obstacle is: lack of solutions that can handle both private and collaborative management of resources together with related Semantic Web data. The thesis presents five requirements for a reusable read/write RDF framework and a concrete software solution that fulfills these requirements. A list of features that have appeared during four major iterations of development is also presented. The third obstacle is: lack of recommendations for how to build learning applications based on Semantic Web technology. The thesis presents seven recommendations in terms of architectures, technologies, frameworks, and type of application to focus on. In addition, as part of the preparatory work to overcome the three obstacles, the thesis also presents a categorization of applications and a derivation of the relations between standards, technologies and application types. / <p>QC 20121105</p>

To develop an assay to detect antibodies to Bacillus thuringiensis subsp. israelensis in mallard ducks, a growth curve was first established for the bacterium. The growth curve indicated that the crystal delta endotoxin would be best harvested from the rest of the cell material after 12 hours of growth. The delta endotoxin was solubilized in alkaline conditions followed by treatment with proteases or no treatment. The two differently treated delta endotoxins were purified by column chromatography. Fractions were assayed for duck erythrocyte lysis and cytotoxicity to a mosquito cell line. The proteolyzed sample gave four protein peaks with gel filtration, and the fourth peak containing biological activity was further separated into three protein fractions by anion exchange chromatography; two of the three showed biological activity. These two fractions contained 22 and 23 kD proteins species. The nonproteolyzed sample was separated into two protein fractions by gel filtration; only the first peak contained the biological activity. This fraction was further separated into two fractions by anion exchange chromatography; only the second fraction, containing a 28 kD protein, exhibited the activity. This fraction contained a 28 kD protein. However, the fractions containing 22 or 23 kD proteins originating from the proteolyzed sample showed the highest biological activity. Mallard ducks were repeatedly exposed to an aerosolized commercial preparation of the organism. Sera were collected periodically and tested for the antibody by an enzyme-linked immunosorbent assay (ELISA). Those toxic antigens containing 22 or 23 kD proteins were unsuitable for the assay. The exposed ducks were found to produce antibodies against the first fraction from anion exchange chromatography of the proteolyzed sample. The antibody titres increased as the number of exposures increased. The results suggest that ELISA is applicable for detecting antibodies against B.t.i. in wild ducks using the fraction containing a 50 kD protein. / Graduation date: 1992

<p>Several variables have been found to affect the Th1/Th2 differentiation of newly activated CD4 T cells. This phenotype can be critical in determining effectiveness of immune responses. Experiments in this thesis were undertaken to better define the in-vivo cellular interactions involved in determining the Th1/Th2 phenotype of newly activated CD4 T cells.</p><p>Lethally irradiated BALB/c mice reconstituted with a constant number of syngeneic, naive spleen cells were challenged with xenogeneic red blood cells (XRBC) conjugated to ovalbumin (OVA) and the Th1/Th2 phenotype of the anti-XRBC response assessed. Antigen-specific interferon-gamma (IFN-g) and interleukin-4 (IL-4) secreting cells obtained from spleens of immunized mice were enumerated by an ELISPOT assay; the relative number of IFN-g- and IL-4-producing cells is taken as a relative measure of Th1 and Th2 components of the response. When challenged with a standard dose of XRBC-OVA, predominant Th1 responses are generated; when challenged with a ten-fold lower dose, such reconstituted mice do not generate significant responses. This adoptive transfer system was employed to explore further the relationships between quantitative changes in the dose of immunizing antigen and the number of responding antigen-specific CD4 T cells, and the Th1/Th2 phenotype of immune responses generated. Unprimed transgenic CD4 T cells specific for OVA can modulate the Th1/Th2 phenotype of the anti-XRBC response upon immunization with XRBC-OVA. Addition of a small number of naive transgenic spleen cells to the standard reconstituting population of normal spleen cells results in the generation of significant numbers of SRBC-specific Th2 cells when mice are challenged with a standard dose, or can generate predominant Th1 responses when mice are challenged with a ten-fold lower dose. Transgenic cells only impact the Th1/Th2 phenotype of CD4 T cells specific for XRBC when OVA is linked to the XRBC. That CD4 T cells specific for different antigens cooperate only through the recognition of linked antigenic determinants has important implications for many aspects of immune regulation. Observations further show that thymocytes from transgenic mice can influence the XRBC-specific response phenotype in an identical manner as transgenic spleen cells, suggesting that previously polarized pro-Th1/Th2 cells are not required in the cooperative events influencing Th1/Th2 phenotype of newly activated CD4 T cells.</p><p>These observations lead to a quantitative description, whereby antigen-mediated CD4 T cell cooperation can affect the Th1/Th2 phenotype of a primary antigen-specific immune response, and provide a context for further analysis at the molecular level.

Graduation date: 1992

<p>Several variables have been found to affect the Th1/Th2 differentiation of newly activated CD4 T cells. This phenotype can be critical in determining effectiveness of immune responses. Experiments in this thesis were undertaken to better define the in-vivo cellular interactions involved in determining the Th1/Th2 phenotype of newly activated CD4 T cells.</p><p>Lethally irradiated BALB/c mice reconstituted with a constant number of syngeneic, naive spleen cells were challenged with xenogeneic red blood cells (XRBC) conjugated to ovalbumin (OVA) and the Th1/Th2 phenotype of the anti-XRBC response assessed. Antigen-specific interferon-gamma (IFN-g) and interleukin-4 (IL-4) secreting cells obtained from spleens of immunized mice were enumerated by an ELISPOT assay; the relative number of IFN-g- and IL-4-producing cells is taken as a relative measure of Th1 and Th2 components of the response. When challenged with a standard dose of XRBC-OVA, predominant Th1 responses are generated; when challenged with a ten-fold lower dose, such reconstituted mice do not generate significant responses. This adoptive transfer system was employed to explore further the relationships between quantitative changes in the dose of immunizing antigen and the number of responding antigen-specific CD4 T cells, and the Th1/Th2 phenotype of immune responses generated. Unprimed transgenic CD4 T cells specific for OVA can modulate the Th1/Th2 phenotype of the anti-XRBC response upon immunization with XRBC-OVA. Addition of a small number of naive transgenic spleen cells to the standard reconstituting population of normal spleen cells results in the generation of significant numbers of SRBC-specific Th2 cells when mice are challenged with a standard dose, or can generate predominant Th1 responses when mice are challenged with a ten-fold lower dose. Transgenic cells only impact the Th1/Th2 phenotype of CD4 T cells specific for XRBC when OVA is linked to the XRBC. That CD4 T cells specific for different antigens cooperate only through the recognition of linked antigenic determinants has important implications for many aspects of immune regulation. Observations further show that thymocytes from transgenic mice can influence the XRBC-specific response phenotype in an identical manner as transgenic spleen cells, suggesting that previously polarized pro-Th1/Th2 cells are not required in the cooperative events influencing Th1/Th2 phenotype of newly activated CD4 T cells.</p><p>These observations lead to a quantitative description, whereby antigen-mediated CD4 T cell cooperation can affect the Th1/Th2 phenotype of a primary antigen-specific immune response, and provide a context for further analysis at the molecular level.

Electrospinning is a technique to achieve nanometer scale fibers. Similar to the conventional spin methods of making fabric, the viscous polymer solution is ejected from a spinneret; stretched and solidified in the air, the solution forms the fibers. The different part of electrospinning among others is that the fibers are driven by the electrostatic force, which induces the repulsion inside the liquid and further reduces the diameter. The resultant product is a non-woven membrane, which is porous; and the pore size is around several nanometers to a micrometer wide. In this work, the relationship between the diameter of electrospun silica fibers, experimental parameters such as concentration and voltage, and between pore size of the fiber membrane and experimental time were studied. Materials used in the process are Polyvinylpyrrolidone (PVP), butanol and spin-on-glass coating solution, which act as polymer carrier, solvent, and silica-precursor, respectively. Polymer/silica precursor composite fibers were ejected from the needle of a plastic syringe when an electrical field, as high as several kV/cm, was applied. Then silica fibers were achieved by baking the composite ones at 773 oK for 12 h. Electrospun silica nanofibers were characterized as a function of polymer solution parameters. The calcined fibers were examined by using a field emission scanning electron microscope. The results showed that the fiber diameters decrease with decreasing proportion of polymer and silica precursor, and increase with a higher electric field. Pore sizes, defined as the grid areas enclosed by fibers on nearby layers, were also examined and showed no time-dependent tendency when the electrospin time was between 1-5 min. Fiber membranes were then used as the platform for protein detection. The results were compared with the control, which used glass slides as the platform. The results make it possible to make a more sensitive biosensing device.