Probiotics in the prevention of clinical manifestations of common infectious diseases in children and in the elderly

Infectious diseases put an enormous burden on both children and the elderly in the forms of respiratory, gastrointestinal and oral infections. There is evidence suggesting that specific probiotics may be antagonistic to pathogens and may enhance the immune system, but the clinical evidence is still too sparce to make general conclusions on the disease-preventive effects of probiotics. This thesis, consisting of four independent, double-blind, placebo-controlled clinical trials, investigated whether Lactobacillus GG (LGG) or a specific probiotic combination containing LGG would reduce the risk of common infections or the prevalence of pathogens in healthy and infection-prone children and in independent and institutionalised elderly people. In healthy day-care children, the 7-month consumption of probiotic milk containing Lactobacillus GG appeared to postpone the first acute respiratory infection (ARI) by one week (p=0.03, adjusted p=0.16), and to reduce complicated infections (39% vs. 47%, p<0.05, adjusted p=0.13), as well as the need for antibiotic treatment (44% vs. 54%, p=0.03, adjusted p=0.08) and day-care absences (4.9 vs. 5.8 days, p=0.03, adjusted p=0.09) compared to the placebo milk. In infection-prone children, the 6-month consumption of a combination of four probiotic bacteria (LGG, L. rhamnosus LC705, Propionibacterium freudenreichii JS, Bifidobacterium breve 99) taken in capsules appeared to reduce recurrent ARIs (72% vs. 82%, p<0.05; adjusted p=0.06), and the effect was particularly noticeable in a subgroup of children with allergic diseases (12% vs. 33%, p=0.03), although no effect on the presence of nasopharyngeal rhinovirus or enterovirus was seen. The 5-month consumption of the same probiotic combination did not show any beneficial effects on the respiratory infections in frail, institutionalised elderly subjects. In healthy children receiving Lactobacillus GG, the reduction in complications resulted in a marginal reduction in the occurrence of acute otitis media (AOM) (31% vs. 39%, p=0.08; adjusted p=0.19), and the postponement of the first AOM episode by 12 days (p=0.04; adjusted p=0.09). However, in otitis-prone children, a probiotic combination did not reduce the occurrence of AOM or the total prevalence of common AOM pathogens (Streptococcus pneumoniae, Haemophilus influenzae, Moraxella catarrhalis), except in the case of children with allergic diseases, in whom probiotics reduced recurrent AOM episodes (0% vs. 14%, p=0.03). In addition, interaction between probiotics and bacterial carriage was seen: probiot-ics reduced AOM in children who did not carry any bacterial pathogens (63% vs. 83%), but the effect was the reverse in children carrying bacteria in the nasopharynx (74% vs 62%) (p<0.05). Long-term probiotic treatment, either LGG given in milk to healthy children for 7 months or a combination of probiotics given in capsules to institutionalised elderly subjects for 5 months, did not reduce the occurrence of acute diarrhoea. However, when the probiotic combination (LGG, L. rhamnosus LC705, Propionibacterium JS) was given in cheese to independent elderly subjects for 4 months, the oral carriage of high Candida counts was reduced in the probiotic group vs. the placebo group (21% vs. 34%, p=0.01, adjusted p=0.004). The risk of hyposalivation was also reduced in the probiotic group (p=0.05). In conclusion, probiotics appear to slightly alleviate the severity of infections by postponing their appearance, by reducing complications and the need for antimicrobial treatments. In addition, they appear to prevent recurrent infections in certain subgroups of children, such as in infection-prone children with allergic diseases. Alleviating ARI by probiotics may lead to a marginal reduction in the occurrence of AOM in healthy children but not in infection-prone children with disturbed nasopharyngeal microbiota. On the basis of these results it could be supposed that Lactobacillus GG or a specific combination containing LGG are effective against viral but not against bacterial otitis, and the mechanism is probably mediated through the stimulation of the immune system. A specific probiotic combination does not reduce respiratory infections in frail elderly subjects. Acute diarrhoea, either in children or in the elderly, is not prevented by the continuous, long-term consumption of probiotics, but the consumption of a specific probiotic combination in a food matrix is beneficial to the oral health of the elderly, through the reduction of the carriage of Candida.

Digital Photography in the Diagnosis and Follow-up of Ocular Diseases

Purpose: The aim of the present study was to develop and test new digital imaging equipment and methods for diagnosis and follow-up of ocular diseases. Methods: The whole material comprised 398 subjects (469 examined eyes), including 241 patients with melanocytic choroidal tumours, 56 patients with melanocytic iris tumours, 42 patients with diabetes, a 52-year old patient with chronic phase of VKH disease, a 30-year old patient with an old blunt eye injury, and 57 normal healthy subjects. Digital 50° (Topcon TRC 50 IA) and 45° (Canon CR6-45NM) fundus cameras, a new handheld digital colour videocamera for eye examinations (MediTell), a new subtraction method using the Topcon Image Net Program (Topcon corporation, Tokyo, Japan), a new method for digital IRT imaging of the iris we developed, and Zeiss photoslitlamp with a digital camera body were used for digital imaging. Results: Digital 50° red-free imaging had a sensitivity of 97.7% and two-field 45° and 50° colour imaging a sensitivity of 88.9-94%. The specificity of the digital 45°-50° imaging modalities was 98.9-100% versus the reference standard and ungradeable images that were 1.2-1.6%. By using the handheld digital colour video camera only, the optic disc and central fundus located inside 20° from the fovea could be recorded with a sensitivity of 6.9% for detection of at least mild NPDR when compared with the reference standard. Comparative use of digital colour, red-free, and red light imaging showed 85.7% sensitivity, 99% specificity, and 98.2 % exact agreement versus the reference standard in differentiation of small choroidal melanoma from pseudomelanoma. The new subtraction method showed growth in four of 94 melanocytic tumours (4.3%) during a mean ±SD follow-up of 23 ± 11 months. The new digital IRT imaging of the iris showed the sphincter muscle and radial contraction folds of Schwalbe in the pupillary zone and radial structural folds of Schwalbe and circular contraction furrows in the ciliary zone of the iris. The 52-year-old patient with a chronic phase of VKH disease showed extensive atrophy and occasional pigment clumps in the iris stroma, detachment of the ciliary body with severe ocular hypotony, and shallow retinal detachment of the posterior pole in both eyes. Infrared transillumination imaging and fluorescein angiographic findings of the iris showed that IR translucence (p=0.53), complete masking of fluorescence (p=0.69), presence of disorganized vessels (p=0.32), and fluorescein leakage (p=1.0) at the site of the lesion did not differentiate an iris nevus from a melanoma. Conclusions: Digital 50° red-free and two-field 50° or 45° colour imaging were suitable for DR screening, whereas the handheld digital video camera did not fulfill the needs of DR screening. Comparative use of digital colour, red-free and red light imaging was a suitable method in the differentiation of small choroidal melanoma from different pseudomelanomas. The subtraction method may reveal early growth of the melanocytic choroidal tumours. Digital IRT imaging may be used to study changes of the stroma and posterior surface of the iris in various diseases of the uvea. It contributed to the revealment of iris atrophy and serous detachment of the ciliary body with ocular hypotony together with the shallow retinal detachment of the posterior pole as new findings of the chronic phase of VKH disease. Infrared translucence and angiographic findings are useful in differential diagnosis of melanocytic iris tumours, but they cannot be used to determine if the lesion is benign or malignant.

Heme oxygenase-1 in cardiovascular diseases

Myocardial infarction (MI) and heart failure are major causes of morbidity and mortality worldwide. Treatment of MI involves early restoration of blood flow to limit infarct size and preserve cardiac function. MI leads to left ventricular remodeling, which may eventually progress to heart failure, despite the established pharmacological treatment of the disease. To improve outcome of MI, new strategies for protecting the myocardium against ischemic injury and enhancing the recovery and repair of the infarcted heart are needed. Heme oxygenase-1 (HO-1) is a stress-responsive and cytoprotective enzyme catalyzing the degradation of heme into the biologically active reaction products biliverdin/bilirubin, carbon monoxide (CO) and free iron. HO-1 plays a key role in maintaining cellular homeostasis by its antiapoptotic, anti-inflammatory, antioxidative and proangiogenic properties. The present study aimed, first, at evaluating the role of HO-1 as a cardioprotective and prohealing enzyme in experimental rat models and at investigating the potential mechanisms mediating the beneficial effects of HO-1 in the heart. The second aim was to evaluate the role of HO-1 in 231 critically ill intensive care unit (ICU) patients by investigating the association of HO-1 polymorphisms and HO-1 plasma concentrations with illness severity, organ dysfunction and mortality throughout the study population and in the subgroup of cardiac patients. We observed in an experimental rat MI model, that HO-1 expression was induced in the infarcted rat hearts, especially in the infarct and infarct border areas. In addition, pre-emptive HO-1 induction and CO donor pretreatment promoted recovery and repair of the infarcted hearts by differential mechanisms. CO promoted vasculogenesis and formation of new cardiomyocytes by activating c-kit+ stem/progenitor cells via hypoxia-inducible factor 1 alpha, stromal cell-derived factor 1 alpha (SDF-1a) and vascular endothelial growth factor B, whereas HO-1 promoted angiogenesis possibly via SDF-1a. Furthermore, HO-1 protected the heart in the early phase of infarct healing by increasing survival and proliferation of cardiomyocytes. The antiapoptotic effect of HO-1 persisted in the late phases of infarct healing. HO-1 also modulated the production of extracellular matrix components and reduced perivascular fibrosis. Some of these beneficial effects of HO-1 were mediated by CO, e.g. the antiapoptotic effect. However, CO may also have adverse effects on the heart, since it increased the expression of extracellular matrix components. In isolated perfused rat hearts, HO-1 induction improved the recovery of postischemic cardiac function and abrogated reperfusion-induced ventricular fibrillation, possibly in part via connexin 43. We found that HO-1 plasma levels were increased in all critically ill patients, including cardiac patients, and were associated with the degree of organ dysfunction and disease severity. HO-1 plasma concentrations were also higher in ICU and hospital nonsurvivors than in survivors, and the maximum HO-1 concentration was an independent predictor of hospital mortality. Patients with the HO-1 -413T/GT(L)/+99C haplotype had lower HO-1 plasma concentrations and lower incidence of multiple organ dysfunction. However, HO-1 polymorphisms were not associated with ICU or hospital mortality. The present study shows that HO-1 is induced in response to stress in both experimental animal models and severely ill patients. HO-1 played an important role in the recovery and repair of infarcted rat hearts. HO-1 induction and CO donor pretreatment enhanced cardiac regeneration after MI, and HO-1 may protect against pathological left ventricular remodeling. Furthermore, HO-1 induction potentially may protect against I/R injury and cardiac dysfunction in isolated rat hearts. In critically ill ICU patients, HO-1 plasma levels correlate with the degree of organ dysfunction, disease severity, and mortality, suggesting that HO-1 may be useful as a marker of disease severity and in the assessment of outcome of critically ill patients.

Real-Time PCR : A Molecular Approach to Investigate the Role of Intestinal Microbiota in the Pathophysiology of Irritable Bowel Syndrome

Irritable bowel syndrome (IBS) is a common multifactorial functional intestinal disorder, the pathogenesis of which is not completely understood. Increasing scientific evidence suggests that microbes are involved in the onset and maintenance of IBS symptoms. The microbiota of the human gastrointestinal (GI) tract constitutes a massive and complex ecosystem consisting mainly of obligate anaerobic microorganisms making the use of culture-based methods demanding and prone to misinterpretation. To overcome these drawbacks, an extensive panel of species- and group-specific assays for an accurate quantification of bacteria from fecal samples with real-time PCR was developed, optimized, and validated. As a result, the target bacteria were detectable at a minimum concentration range of approximately 10 000 bacterial genomes per gram of fecal sample, which corresponds to the sensitivity to detect 0.000001% subpopulations of the total fecal microbiota. The real-time PCR panel covering both commensal and pathogenic microorganisms was assessed to compare the intestinal microbiota of patients suffering from IBS with a healthy control group devoid of GI symptoms. Both the IBS and control groups showed considerable individual variation in gut microbiota composition. Sorting of the IBS patients according to the symptom subtypes (diarrhea, constipation, and alternating predominant type) revealed that lower amounts of Lactobacillus spp. were present in the samples of diarrhea predominant IBS patients, whereas constipation predominant IBS patients carried increased amounts of Veillonella spp. In the screening of intestinal pathogens, 17% of IBS samples tested positive for Staphylococcus aureus, whereas no positive cases were discovered among healthy controls. Furthermore, the methodology was applied to monitor the effects of a multispecies probiotic supplementation on GI microbiota of IBS sufferers. In the placebo-controlled double-blind probiotic intervention trial of IBS patients, each supplemented probiotic strain was detected in fecal samples. Intestinal microbiota remained stable during the trial, except for Bifidobacterium spp., which increased in the placebo group and decreased in the probiotic group. The combination of assays developed and applied in this thesis has an overall coverage of 300-400 known bacterial species, along with the number of yet unknown phylotypes. Hence, it provides good means for studying the intestinal microbiota, irrespective of the intestinal condition and health status. In particular, it allows screening and identification of microbes putatively associated with IBS. The alterations in the gut microbiota discovered here support the hypothesis that microbes are likely to contribute to the pathophysiology of IBS. The central question is whether the microbiota changes described represent the cause for, rather than the effect of, disturbed gut physiology. Therefore, more studies are needed to determine the role and importance of individual microbial species or groups in IBS. In addition, it is essential that the microbial alterations observed in this study will be confirmed using a larger set of IBS samples of different subtypes, preferably from various geographical locations.

Cyclin dependent protein kinases as drug targets – potential in cardiovascular diseases

Complications of atherosclerosis such as myocardial infarction and stroke are the primary cause of death in Western societies. The development of atherosclerotic lesions is a complex process, including endothelial cell dysfunction, inflammation, extracellular matrix alteration and vascular smooth muscle cell (VSMC) proliferation and migration. Various cell cycle regulatory proteins control VSMC proliferation. Protein kinases called cyclin dependent kinases (CDKs) play a major role in regulation of cell cycle progression. At specific phases of the cell cycle, CDKs pair with cyclins to become catalytically active and phosphorylate numerous substrates contributing to cell cycle progression. CDKs are also regulated by cyclin dependent kinase inhibitors, activating and inhibitory phosphorylation, proteolysis and transcription factors. This tight regulation of cell cycle is essential; thus its deregulation is connected to the development of cancer and other proliferative disorders such as atherosclerosis and restenosis as well as neurodegenerative diseases. Proteins of the cell cycle provide potential and attractive targets for drug development. Consequently, various low molecular weight CDK inhibitors have been identified and are in clinical development. Tylophorine is a phenanthroindolizidine alkaloid, which has been shown to inhibit the growth of several human cancer cell lines. It was used in Ayurvedic medicine to treat inflammatory disorders. The aim of this study was to investigate the effect of tylophorine on human umbilical vein smooth muscle cell (HUVSMC) proliferation, cell cycle progression and the expression of various cell cycle regulatory proteins in order to confirm the findings made with tylophorine in rat cells. We used several methods to determine our hypothesis, including cell proliferation assay, western blot and flow cytometric cell cycle distribution analysis. We demonstrated by cell proliferation assay that tylophorine inhibits HUVSMC proliferation dose-dependently with an IC50 value of 164 nM ± 50. Western blot analysis was used to determine the effect of tylophorine on expression of cell cycle regulatory proteins. Tylophorine downregulates cyclin D1 and p21 expression levels. The results of tylophorine’s effect on phosphorylation sites of p53 were not consistent. More sensitive methods are required in order to completely determine this effect. We used flow cytometric cell cycle analysis to investigate whether tylophorine interferes with cell cycle progression and arrests cells in a specific cell cycle phase. Tylophorine was shown to induce the accumulation of asynchronized HUVSMCs in S phase. Tylophorine has a significant effect on cell cycle, but its role as cell cycle regulator in treatment of vascular proliferative diseases and cancer requires more experiments in vitro and in vivo.

Molekyylibiologisten menetelmien kehittäminen ihmisen suoliston sulfaattia pelkistävien bakteerien määrittämiseksi

Ihmisen ruuansulatuskanavan bakteeriston kehitys alkaa syntymästä, jolloin ensimmäiset bakteerit kansoittavat steriilin ruuansulatuskanavan. Bakteeristo kehittyy perimän, ympäristön ja varhaisen ruokavalion vaikutuksesta kohti monimuotoisempaa bakteeripopulaatiota. Aikuisen ruuansulatuskanavan normaalibakteeristo on varsin muuttumaton, mutta siihen vaikuttavat monet tekijät, kuten ikä, terveydentila, ruokavalio ja antibioottien käyttö. Bakteeriston koostumus vaihtelee ruuansulatuskanavan eri osissa ja bakteerimäärä kasvaa kohti paksusuolta, ollen paksusuolessa ja ulosteessa peräti 1010-1012 pmy/ml. Suurin osa ruuansulatuskanavan bakteereista on anaerobeja. Ruuansulatuskanavan bakteeristo vaikuttaa muun muassa suoliston kehittymiseen ja hiilihydraattien ja proteiinien hajotukseen sekä toimii osana immuunipuolustusta. Sulfaattia pelkistävät bakteerit (SRB) ovat monimuotoinen ryhmä pääosin anaerobisia bakteereita, jotka käyttävät aineenvaihdunnassaan elektronin vastaanottajana sulfaattia muuttaen sen lopulta sulfidiksi. SRB:t ovat sopeutuneet useisiin erilaisiin ympäristöihin. Niitä tavataan mm. vesistöjen sedimenteissä sekä ihmisen ruuansulatuskanavassa. Ihmisen ruuansulatuskanavassa on SRB:ta n. 105-108 pmy/g, ja niitä on löydetty erityisesti anaerobisista osista kuten suun ientaskuista ja paksusuolesta. SRB:t voivat olla haitaksi ruuansulatuskanavalle tuottamansa sulfidin vuoksi, joka esiintyy vesiliuoksessa vetysulfidina. Tämän on havaittu olevan toksista suoliston epiteelisoluille. Viimeaikoina on kiinnostuttu sulfaatinpelkistäjien yhteydestä suoliston sairaustiloihin, kuten tulehduksellisiin suolistosairauksiin (IBD). Pro gradu -tutkimukseni tavoitteena oli kehittää PCR-DGGE- ja qPCR-menetelmät ulosteen sulfaattia pelkistävien bakteerien määritykseen. Kohdegeeninä menetelmänkehityksessä käytettiin dsrAB-geeniä, joka koodaa dissimilatorista sulfiitinpelkistysentsyymiä. dsrAB-geeni on sulfaatinpelkistäjille ominainen konservoitunut geenialue, johon perustuvia tutkimuksia ei vielä ole paljon ihmispuolelta. qPCR-menetelmä saatiin optimoitua herkäksi ja spesifiseksi käyttäen dsrA-geenispesifisiä alukkeita, mutta PCR-DGGE-menetelmää ei saatu optimoitua käytössä olleilla alukkeilla, jotka monistivat PCR-DGGE:ssa myös negatiivikontrollikantoja. Tutkittaessa qPCR:lla IBD:tä (Crohn ja ulseratiivinen koliitti) sairastavien lasten ja terveiden kontrollihenkilöiden ulostenäytteistä eristettyä DNA:ta, merkittävää eroa SRB-määrissä ei havaittu eri ryhmien välillä. Crohnin tautia sairastavien aktiivisen vaiheen ja oireettoman vaiheen näytteiden välillä oli kuitenkin tilastollisesti merkitsevä ero (SRB-määrät; oireeton vaihe>oireellinen vaihe) (P <0,05).

Lower Gastrointestinal Microbiota in Health and Irritable Bowel Syndrome : Characterisation and Effect of Probiotic Intervention

The human gastrointestinal (GI) microbiota is a complex ecosystem that lives in symbiosis with its host. The growing awareness of the importance of the microbiota to the host as well as the development of culture-free laboratory techniques and computational methods has enormously expanded our knowledge of this microbial community. Irritable bowel syndrome (IBS) is a common functional bowel disorder affecting up to a fifth of the Western population. To date, IBS diagnosis has been based on GI symptoms and the exclusion of organic diseases. The GI microbiota has been found to be altered in this syndrome and probiotics can alleviate the symptoms, although clear links between the symptoms and the microbiota have not been demonstrated. The aim of the present work was to characterise IBS related alterations in the intestinal microbiota, their relation to IBS symptoms and their responsiveness to probiotic theraphy. In this thesis research, the healthy human microbiota was characterised by cloning and sequencing 16S rRNA genes from a faecal microbial community DNA pool that was first profiled and fractionated according to its guanine and cytosine content (%G+C). The most noticeable finding was that the high G+C Gram-positive bacteria (the phylum Actinobacteria) were more abundant compared to a corresponding library constructed from the unfractionated DNA pool sample. Previous molecular analyses of the gut microbiota have also shown comparatively low amounts of high G+C bacteria. Furthermore, the %G+C profiling approach was applied to a sample constructed of faecal DNA from diarrhea-predominant IBS (IBS-D) subjects. The phylogenetic microbial community comparison performed for healthy and IBS-D sequence libraries revealed that the IBS-D sample was rich in representatives of the phyla Firmicutes and Proteobacteria whereas Actinobacteria and Bacteroidetes were abundant in the healthy subjects. The family Lachnospiraceae within the Firmicutes was especially prevalent in the IBS-D sample. Moreover, associations of the GI microbiota with intestinal symptoms and the quality of life (QOL) were investigated, as well as the effect of probiotics on these factors. The microbial targets that were analysed with the quantitative real-time polymerase chain reaction (qPCR) in this study were phylotypes (species definition according to 16S rRNA gene sequence similarity) previously associated with either health or IBS. With a set of samples, the presence or abundance of a phylotype that had 94% 16S rRNA gene sequence similarity to Ruminococcus torques (R. torques 94%) was shown to be associated with the severity of IBS symptoms. The qPCR analyses for selected phylotypes were also applied to samples from a six-month probiotic intervention with a mixture of Lactobacillus rhamnosus GG, L. rhamnosus Lc705, Propionibacterium freudenreichii ssp. shermanii JS and Bifidobacterium breve Bb99. The intervention had been previously reported to alleviate IBS symptoms, but no associations with the analysed microbiota representatives were shown. However, with the phylotype-specific assays applied here, the abundance of the R. torques 94% -phylotype was shown to be lowered in the probiotic-receiving group during the probiotic supplementation, whereas a Clostridium thermosuccinogenes 85% phylotype, previously associated with a healthy microbiota, was found to be increased compared to the placebo group. To conclude, with the combination of methods applied, higher abundance of Actinobacteria was detected in the healthy gut than found in previous studies, and significant phylum-level microbiota alterations could be shown in IBS-D. Thus, the results of this study provide a detailed overview of the human GI microbiota in healthy subjects and in subjects with IBS. Furthermore, the IBS symptoms were linked to a particular clostridial phylotype, and probiotic supplementation was demonstrated to alter the GI microbiota towards a healthier state with regard to this and an additional bacterial phylotype. For the first time, distinct phylotype-level alterations in the microbiota were linked to IBS symptoms and shown to respond to probiotic therapy.

Determination of intestinal permeability in healthy horses using the contrast medium iohexol

Tämä lisensiaatin tutkielma koostuu kolmesta osasta; kirjallisuuskatsauksesta, kokeellisesta osasta ja liitteistä. Iohexol on ionisoitumaton, trijodattu ja vesiliukoinen röntgenvarjoaine. Iohexolia on hyödynnetty lääketieteessä useita vuosia. Iohexolia on käytetty muun muassa angio- ja myelografiassa, lisäksi iohexolia on hyödynnetty arvioitaessa munuaiskerästen suodattumisnopeutta sekä suoliston läpäisevyyden muutoksia. Hevosen tulehduksellisessa suolistosairaudessa (Inflammatory bowel disease, IBD) suoliston rakenne ja sen läpäisevyys muuttuu; tyypillistä on tulehdussolujen kertyminen suoliston seinämään ja myös sidekudosmuodostusta saattaa esiintyä. Suolisto muutoksia saatetaan havaita sekä ohut- että paksusuolessa. IBD aiheuttaa hevoselle laihtumista, johtuen ravintoaineiden puutteellisesta imeytymisestä ja proteiinien menetyksestä suoleen suoliston häiriötilan yhteydessä. Tällä hetkellä IBD:n diagnostiikka perustuu tyypillisiin oireisiin, kliiniseen tutkimukseen, verinäytteisiin, glukoosin imeytymistestiin ja peräsuolesta otettuun koepalaan. IBD:n diagnostiikka on kuitenkin erittäin haastavaa ja tutkimusmenetelmiin liittyy lukuisia ongelmia, jotka vähentävät niiden luotettavuutta IBD:n diagnostiikassa. Tutkimuksemme tarkoituksena on kehittää hevosen IBD:n diagnostiikkaa entistä helpompaan, luotettavampaan ja turvallisempaan suuntaan. Tämän alustavan tutkimuksen tavoitteet olivat: (1) tutkia voidaanko iohexol havaita hevosen seerumissa oraalisen annostelun jälkeen ja (2) muodostaa iohexolin pitoisuuskuvaaja ajan funktiona terveillä hevosilla. Materiaalimme koostui kymmenestä terveestä hevosesta, joilla ei ollut havaittu laihtumista tai ripulia. Ennen iohexolin annostelua hevosille suoritettiin kliininen tutkimus ja verinäytteet otettiin maha-suolikanavan sairauden poissulkemiseksi. Hevosille suoritettiin myös mahalaukun tähystys. 16 tunnin paaston jälkeen 1 ml/kg Iohexolia annosteltiin 10 % -liuoksena nenämahaletkulla suoraan mahaan ja verinäytteet otettiin 0, 30, 60, 120, 180, 240, 300 ja 360 minuuttia annostelun jälkeen. Iohexolin pitoisuus määritettiin käyttämällä korkean erotuskyvyn nestekromatografiaa. Iohexolin pitoisuuksista tietyillä ajanhetkillä muodostettiin kuvaaja. Hevosilla ei havaittu maha-suolikanavan sairauksia. Kaikki hevoset olivat hyvässä kuntoluokassa ja mahalaukun tähystyksessä ei havaittu merkittäviä muutoksia. Verinäytteiden tulokset olivat viiterajoissa. Kaikki hevoset sietivät iohexolia hyvin ja haittavaikutuksia ei havaittu. Iohexol oli havaittavissa seerumissa 60 minuutin kuluttua annostelusta. Kuvaajassa voitiin havaita kaksi huippua. Statistiset menetelmät tukivat löydöksiä. Iohexol testi oli yksinkertainen suorittaa ja siihen ei liittynyt haittavaikutuksia. Annos 1ml/kg oli havaittavissa seerumissa. Iohexolin pitoisuuskuvaaja muodosti kaksi huippua, ja tämänkaltainen ilmiö on kuvattu kirjallisuudessa aikaisemmin useiden lääkkeiden tapauksessa. Hevosella ilmiö liittyy todennäköisesti maha-suolikanavan rakenteellisiin ja fysiologisiin eroavaisuuksiin ja lisätutkimuksia ilmiön varmistamiseksi tarvitaan. Iohexol näyttää olevan potentiaalinen merkkiaine suoliston läpäisevyyden arviointiin ja lisätutkimuksia IBD:tä sairastavien hevosten seerumin iohexolin pitoisuuksista verrattuna terveiden hevosten seerumin iohexolin pitoisuuksiin on suunnitteilla.