Large-scale serum analysis identifies unique systemic biomarkers in psoriasis and hidradenitis suppurativa.

BACKGROUND: Hidradenitis suppurativa (HS) is now recognized as a systemic inflammatory disease, sharing molecular similarities with psoriasis. Direct comparison of the systemic inflammation in HS with psoriasis is lacking. OBJECTIVES: To evaluate the serum proteome of HS and psoriasis, and to identify biomarkers associated with disease severity. METHODS: In this cross-sectional study, 1536 serum proteins were assessed using the Olink Explore (Proximity Extension Assay) high-throughput panel in patients with moderate-to-severe HS (n = 11), patients with psoriasis (n = 10) and age- and body mass index-matched healthy controls (n = 10). RESULTS: HS displayed an overall greater dysregulation of circulating proteins, with 434 differentially expressed proteins (absolute fold change ≥ 1·2; P ≤ 0·05) in patients with HS vs. controls, 138 in patients with psoriasis vs. controls and 503 between patients with HS and patients with psoriasis. Interleukin (IL)-17A levels and T helper (Th)1/Th17 pathway enrichment were comparable between diseases, while HS presented greater tumour necrosis factor- and IL-1ß-related signalling. The Th17-associated markers peptidase inhibitor 3 (PI3) and lipocalin 2 (LCN2) were able to differentiate psoriasis from HS accurately. Both diseases presented increases of atherosclerosis-related proteins. Robust correlations between clinical severity scores and immune and atherosclerosis-related proteins were observed across both diseases. CONCLUSIONS: HS and psoriasis share significant Th1/Th17 enrichment and upregulation of atherosclerosis-related proteins. Despite the greater body surface area involved in psoriasis, HS presents a greater serum inflammatory burden.

Doppler ultrasound-based noninvasive biomarkers in hidradenitis suppurativa: evaluation of analytical and clinical validity.

BACKGROUND: There is a need for valid and reliable biomarkers in hidradenitis suppurativa (HS) for diagnosis and disease activity monitoring. Imaging-based biomarkers have the potential to fulfil this unmet need but no evaluation of analytical or clinical validity has yet been undertaken. OBJECTIVES: To evaluate the analytical and clinical validity of sonographic epidermal thickness, Doppler ultrasound and dermal tunnel diameter in patients with HS. METHODS: Twenty-two participants with HS were recruited and underwent a total of 65 matched ultrasound and skin biopsies of lesional, perilesional and unaffected tissue. Ultrasound measurements were performed in triplicate with mean values used. Skin biopsies underwent immunohistochemistry as per previously published methods. Analytical validity was assessed in individual ultrasound-biopsy pairs (n = 65) by comparisons of sonographic variables with histological correlates. Clinical validity was assessed in individual patients (n = 22) by comparing measures of overall disease activity with sonographic outcomes. RESULTS: Epidermal thickness, dermal tunnel diameter and power Doppler intensity were assessed. Sonographic epidermal thickness and dermal tunnel diameter have high analytical validity with corresponding histological measurements. Power Doppler intensity demonstrated high correlation with dermal CD3+ and CD11c+ cell counts but not neutrophil elastase-positive cells. Power Doppler ultrasound has significant correlation with pain scores, abscess and nodule count, International HS Severity Scoring System score and number of draining tunnels. CONCLUSIONS: Sonographic epidermal thickness and dermal tunnel diameter have acceptable levels of analytical validity in the assessment of HS lesions. Power Doppler intensity demonstrates acceptable clinical and analytical validity, suggesting it is a valid imaging-based biomarker in HS.

Dermal tunnels influence time to clinical response and family history influences time to loss of clinical response in patients with hidradenitis suppurativa treated with adalimumab.

BACKGROUND: Clinical response in hidradenitis suppurativa (HS) is most commonly assessed using the Hidradenitis Suppurativa Clinical Response (HiSCR) measure. Dermal tunnels, increased body mass index, smoking and antibiotic use significantly decrease the odds of achieving HiSCR. However, there are few data exploring if clinical features are also associated with length of time to achieve clinical response and/or time to lose clinical response. AIM: To explore whether variables associated with achievement of HiSCR are associated with time to achieve HiSCR and time to loss of HiSCR in patients with HS treated with adalimumab 40 mg weekly in the PIONEER open-label extension study. METHODS: Time-to-event analyses were performed to estimate time to achieve HiSCR and time to loss of HiSCR. The log rank test was used to compare cumulative incidence curves for a priori patient- and disease-associated factors. Cox regression analysis was performed to compare time-to-event outcomes in the presence of a priori variables. All statistical analyses were completed with R software (V3.5.3). RESULTS: Presence of dermal tunnels significantly increased the time to achieve HiSCR (median 32.6 vs. 14.3 weeks, P = 0.02) and the hazard ratio (HR) was significant after controlling for patient and disease factors (HR = 0.70, 95% CI 0.51-0.96, P = 0.03). A positive family history of HS significantly decreased the time to loss of HiSCR (median 11.4 vs. 18 weeks, P < 0.001) and remained significant in Cox regression analysis (HR = 2.01, 95% CI 1.40-2.88, P < 0.001). CONCLUSION: The presence of dermal tunnels significantly influences the odds of achieving HiSCR and the time to achieve HiSCR, while family history influences time to loss of HiSCR.

Malignancy and infection risk during adalimumab therapy in hidradenitis suppurativa.

BACKGROUND: The association of adalimumab therapy with malignancy and infection is established in other inflammatory diseases; however, rates of hidradenitis suppurativa (HS) are based on case reports or retrospective healthcare data and the effect of adalimumab therapy on these rates is unknown. Previously reported rates in the PIONEER OLE Phase 3 study reported on rates only in a subpopulation of 88 participants rather than the entire cohort. AIM: To quantify rates of malignancy and serious infection in all patients with HS treated with adalimumab 40 mg weekly. METHODS: Reanalysis was undertaken of individual patient data from the PIONEER 1, PIONEER 2 and PIONEER open-label extension Phase 3 trial data encompassing 591 unique patients with HS administered adalimumab 40 mg weekly without concurrent antibiotic exposure. Incidence rates of serious infection and malignancy were calculated. RESULTS: Incidence rates of serious infection and malignancy were 2.14 and 0.46 per 100 patient-years, respectively. Rates of infection and malignancy were comparable to those in other inflammatory conditions examined. CONCLUSION: Incidence of serious infection in patients with HS on adalimumab is comparable to those with psoriasis and inflammatory arthropathies, but the incidence of malignancy is increased. This may reflect disease-specific malignancy risk rather than an effect of adalimumab.

Contagious agalactia monitoring in caprine herds through regular bulk tank milk sampling.

Surveillance and control of Mycoplasma spp. responsible for contagious agalactia (CA) in caprine herds are important challenges in countries with a large small-ruminant dairy industry. In the absence of any clinical signs, being able to determine the potential circulation of mycoplasmas within a herd could help to prevent biosecurity issues during animal exchanges between farms and improve health management practices. The objective of this study was to determine whether regular sampling of bulk tank milk was suitable for such surveillance. Twenty farms were sampled once a month for 2 yr and CA-responsible mycoplasmas were detected by real-time PCR on DNA extracted from milk, using 3 different DNA extraction methods. The pattern of mycoplasma excretion in bulk tank milk was assessed over time and several herd characteristics were recorded together with any event occurring within the herds. In general, the results obtained with the different detection methods were comparable and mainly agreed with the culture results. Several patterns of excretion were observed but were not related to herd characteristics (size, breed, and so on). Recurrence of the same (sub)species and same pulsed-field gel electrophoresis subtype during the 2-yr period is indicative of the considerable persistence of mycoplasmas. This persistence was associated with intermittent excretion. In conclusion, bulk tank milk sampling could be valuable for controlling CA in caprine herds provided it is repeated several times, yet to be defined, per year and analyzed using an appropriate methodology and the right cut-off for interpretation.

Opportunistic screening for bone disease using abdominal CT scans obtained for other reasons in newly diagnosed IBD patients.

Bone disease is prevalent among patients with inflammatory bowel disease (IBD), though bone density screening remains underutilized. We used CT scans performed for other indications in IBD patients to identify and monitor osteopenia using CT attenuation values at the lumbar spine. Significant rates of bone disease were detected which would have otherwise gone undiagnosed. INTRODUCTION: Osteoporosis affects about 14-42% of patients with IBD. Though screening is recommended in IBD patients with risk factors, it remains underutilized. In patients with newly diagnosed IBD, we used CT scans performed for other indications to identify and monitor progression of osteopenia. METHODS: Using the Ocean State Crohn's and Colitis Area Registry, we identified adult patients with one or more abdominal CT scans. Each patient had two age- and gender-matched controls. Radiologists measured attenuation through trabecular bone in the L1 vertebral body recorded in Hounsfield units (HU). Generalized estimating equations were used to measure how HU varied as a function of gender, type of IBD, and age. RESULTS: One hundred five IBD patients were included, and 72.4% were classified as "normal" bone mineral density (BMD) and 27.6% as potentially osteopenic: 8.6% with ulcerative colitis and 19.0% with Crohn's disease. We found a decrease in bone density over time (p < 0.001) and that BMD decreases more in Crohn's disease than in ulcerative colitis (p < 0.004). Sixty patients had two CT scans, and mean loss of 9.3 HU was noted. There was a non-significant decrease in BMD over time in patients exposed to > 31 days of steroids and BMD was stable with < 30 days of steroid exposure (p < 0.09). CONCLUSION: Using CT scans obtained for other indications, we found low rates of osteopenia and osteoporosis that may otherwise have gone undiagnosed. Refinement of opportunistic screening may have advantages in terms of cost-savings and earlier detection of bone loss.

Scan length optimization for pulmonary embolism at CT angiography: analysis based on the three-dimensional spatial distribution of 370 emboli in 100 patients.

AIM: To determine the size and three-dimensional spatial distribution of pulmonary emboli (PE) at computed tomography angiography (CTA) to optimize the scan length. MATERIALS AND METHODS: Two experienced radiologists jointly reviewed 100 consecutive, positive PE CTA studies performed in the Emergency Department (53 women; age 61±17 years). All studies were conducted on a 16-detector row CT machine. In each case, the number of emboli was counted and the proximal and distal spatial coordinates of each embolus documented. Coordinates of the main pulmonary artery bifurcation (MPAb) and carina were recorded. For normalization, the thoracic cavity height (H)-from inlet to lowest hemidiaphragm-was measured. The minimal scan lengths for (a) capturing all emboli and (b) rendering a positive diagnosis were determined. RESULTS: Three hundred and seventy (370) emboli were detected. The average number of PE per patient was 3.7 (maximum 12, minimum 1). Their average length was 2.7 cm. Nine patients had saddle emboli (9%), and 71% of emboli were at or below the MPAb. An 18 cm (0.90×H) scan length, centred 4 cm (0.18×H) below the carina, captures all PE in this dataset while reducing z-axis coverage by 29% (34% for normalized data). Moreover, a 14.2 cm (0.78×H) scan length appropriately centred captures at least one embolus in all patients while reducing coverage by 44% (43%). Decreasing scan length to the lesser of 14.2 cm and 0.78×H per patient reduces coverage by 47%. CONCLUSION: Scan length at CTA for PE can be reduced by up to 47% while preserving diagnostic accuracy for PE detection.

Thymic and extrathymic origins of gut intraepithelial lymphocyte populations in mice.

We have investigated the origin of intraepithelial lymphocytes (IEL) populations in the murine gut, using reconstitution experiments in which the presence of thymus-derived cells of host or donor origin is rigorously controlled: RAG-/- mutant mice which have no T cells, were injected either with the bone marrow (BM) cells of nude mice or with selected peripheral lymph node (LN) T cells of euthymic mice. In thymectomized RAG-/- mice, injection of BM cells from nude mice led, after 2 mo, to the development of a peripheral B cell compartment and to the appearance, in the gut, of IEL bearing homodimeric CD8 alpha chains and either gamma/delta or alpha/beta TCR. In RAG-/- mice with a thymus, a similar injection led to complete lymphoid reconstitution, with the additional appearance in the gut of CD4+, CD8 alpha/beta+ or CD4+CD8 alpha/alpha+ IEL, all bearing alpha/beta TCR. In contrast, injection of LN T cells into these mice reconstituted a gut IEL population made of CD4+, CD8 alpha/beta+, or CD4+ CD8 alpha/alpha+ cells, all bearing alpha/beta TCR; CD8 alpha/alpha+ TCR-gamma/delta+ or alpha/beta+ IEL were not observed. These results demonstrate that the thymus and/or thymic-derived peripheral T cells are absolutely required for the generation of CD4+, CD8 alpha/beta+, and CD4+CD8 alpha/alpha+ IEL, which are thus thymus dependent. In contrast, TCR+ CD8 alpha/alpha+ IEL appear in the absence of the thymus, and thus are thymus independent.

The V beta repertoire of mouse gut homodimeric alpha CD8+ intraepithelial T cell receptor alpha/beta + lymphocytes reveals a major extrathymic pathway of T cell differentiation.

Gut intraepithelial lymphocytes (IEL) contain two independent T cell receptor alpha/beta + T cell populations, with different V beta repertoires. In DBA/2 mice (Mlsa, IE+), the CD4+ and heterodimeric alpha/beta CD8+ thymodependent T cell pool shows the same deletion of V beta 6, 8.1, and 11+ cells as found in peripheral lymphoid organs. In contrast, such deletions are not observed in the pool of IEL bearing homodimeric alpha CD8+ chains, in which these V beta families are frequently observed in high amounts. The size of this gut homodimeric alpha CD8+ IEL pool and its different V beta repertoire selection demonstrate the existence of a major extrathymic pathway of T cell differentiation with a gut-restricted localization. The large amount of the thymo-independent, homodimeric alpha CD8+ IEL found in the small bowel may contribute to a first line of defense against exogenous superantigens.

The mouse gut T lymphocyte, a novel type of T cell. Nature, origin, and traffic in mice in normal and graft-versus-host conditions.

Lymphocytes of the mouse intestinal mucosa, identified in tissue sections or purified suspensions of intraepithelial lymphocytes as T cells (gut T lymphocytes [GTL]), were studied in normal mice or in beige mice (the equivalent of the Chediak-Higashi syndrome in man, characterized by giant granules in various cell types, including mast cells). Mice were studied in normal or in germ-free conditions, or during a graft versus host (GVH) reaction resulting from the injection of parental thymocytes into lethally irradiated F1 mice, a condition leading to massive accumulation of T lymphocytes of donor origin in the host gut mucosa. In normal as well as in GVH conditions, a high percentage of the gut IE lymphocytes contain granules (up to 80% in the beige mouse). These granules have ultrastructural, hostochemical and other features resembling those of mast cell granules; in beige mice, up to 50% of them can be shown to contain histamine. Granulated T cells are also found in the lamina propria. It appears that the GTL may progressively lose their surface T antigens when the granules become more developed. Kinetics of [3H]TdR labeling of the GTL, transfer experiments with T cells of various origins, selective [3H]TdR labeling and selective irradiation of the Peyer's patches (PP), and effect of thoraic duct (TD) drainage led to the conclusion that GTL are the progeny of T cells stimulated to divide in the PP microenvironment, which endows them with a gut-homing tendency. From the PP, these cells follow a cycle, migrating to the TD and to the blood to colonize the whole intestinal mucosa, the majority of them as dividing cells undergoing a single round of traffic, with some probably able to recirculate and becoming a more long-lived variety. Antigenic stimulation within the PP is necessary for the emergence of GTL progenitors, but their gut-homing property is unrelated to the antigen as shown with fetal gut grafts, notably in GVH where grafts syngeneic to the host or donor become similarly infiltrated by GTL. On the basis of their properties and of further evidence to be reported elsewhere, it is proposed that GTL belong to a special class of T lymphocytes, related to the immune defenses of the mucosal systems in general, and capable of acting as progenitors of mucosal mast cells.

Cytotoxic differentiation of mouse gut thymodependent and independent intraepithelial T lymphocytes is induced locally. Correlation between functional assays, presence of perforin and granzyme transcripts, and cytoplasmic granules.

Mouse gut intraepithelial lymphocytes (IEL), whether thymodependent (CD4+ or CD8 alpha/beta +; TCR-alpha/beta +) or thymoindependent (CD8 alpha/alpha +; TCR-alpha/beta + or -gamma/delta +), all display cytotoxic activity in a "redirected lysis assay" using anti-CD3 or anti-TCR beta or delta chains secreting hybridomas as targets; this is also observed with IEL of germ-free mice, indicating that this activity, which is absent in peripheral T lymphocytes, does not require stimulation by bacterial antigens. Perforin and granzyme transcripts are detectable in unselected gut IEL, in contrast to normal T lymphocytes of peripheral lymphoid organs. Cytological labeling (with [3H]DFP) of IEL smears reveals labeled granules (i.e., containing serine-esterases, presumably granzymes) in all subsets of gut IEL. This indicates that the gut micro-environment has an inductive role on the cytotoxic differentiation of lymphocytes of various origins when they reach the gut wall to become IEL.

Oligoclonal repertoire of the CD8 alpha alpha and the CD8 alpha beta TCR-alpha/beta murine intestinal intraepithelial T lymphocytes: evidence for the random emergence of T cells.

The epithelium of the small intestine in normal euthymic mice contains a large number of intraepithelial lymphocytes (IEL), some of which bear a T cell receptor alpha/beta (TCR-alpha/beta). About half of these TCR-alpha/beta IEL display the CD8 alpha alpha phenotype and the remaining have the CD8 alpha beta or the CD4 phenotypes. To examine whether TCR-alpha/beta IEL have a TCR-beta chain repertoire as diverse as that of TCR-alpha/beta lymph node lymphocytes (LNL), we used a recently described PCR technique that allows a global analysis of the TCR-beta chain repertoire. Within any given mouse, the repertoires expressed in both CD8 alpha alpha and CD8 alpha beta TCR-alpha/beta IEL populations are oligoclonal and nonoverlapping between the two subsets. The clones are largely conserved through the length of the small intestine of the same individual. However, genetically identical individuals raised under indistinguishable environmental conditions display distinct oligoclonal repertoires. Those findings indicate that few cells of CD8 alpha alpha or of the CD8 alpha beta phenotype are responsible for the repopulation of the intestinal epithelium.

Gut mucosal mast cells. Origin, traffic, and differentiation.

Gut mucosal mast cells (MMC), which are nearly absent in normal mice are abundant during nematode infection. In normal mice, study of MMC precursors (MMC-P: cells giving rise to MMC colonies in the presence of IL-3) show that: (a) their frequency, judged by limiting dilution is very high in bone marrow (BM) and gut, and very low in most lymphoid organs and thoracic duct lymph (TDL); (b) gut MMC-P are Thy-1- Lyt-1-2- and are not rapidly replicating; (c) they are the progeny of less differentiated BM MMC-P which are attracted from the blood to the gut mucosa by local factor(s), other than antigen and T cell factors (since normal amounts of gut MMC-P are found in germ-free, nude, and newborn mice). In mice bearing the Wehi 3 tumor (which releases enough IL-3 to produce detectable blood levels) spleen and mesenteric lymph nodes (LN) show increased MMC-P frequency, the greatest increase being in the gut and BM, where numerous differentiated MMC are found. In Nippostrongylus brasiliensis (Nb)-infested mice (known to develop a large, T cell-dependent, gut MMC infiltration), gut MMC-P proliferation is induced by IL-3 released from gut mucosal Thy-1+ Lyt-2- cells, whose in vitro IL-3 release capability is much higher than that of similar cells from normal mice. Both Nb-stimulated T blasts and proliferating MMC-P undergo cyclic traffic, migrating into the TDL and then seeding the whole length of the gut (a process which allows a widespread immune defense after a local antigenic stimulus). Experiments using 2-d interruption of this traffic and fetal gut grafts, suggest that the continuous homing of T blasts back to the gut which leads to permanent Nb-stimulated IL-3 release, is essential for the full maturation of MMC. Transfer experiments in the rat show that TDL circulating MMC-P rapidly mature into MMC when they home back to the Nb-infested gut. It is proposed that gut MMC arise after several stages of progressive differentiation of MMC-P, influenced both by IL-3 and unidentified gut factor(s).

Critical role for the common cytokine receptor gamma chain in intrathymic and peripheral T cell selection.

The common cytokine receptor gamma chain (gammac), which is a functional subunit of the receptors for interleukins (IL)-2, -4, -7, -9, and -15, plays an important role in lymphoid development. Inactivation of this molecule in mice leads to abnormal T cell lymphopoiesis characterized by thymic hypoplasia and reduced numbers of peripheral T cells. To determine whether T cell development in the absence of gammac is associated with alterations of intrathymic and peripheral T cell selection, we have analyzed gammac-deficient mice made transgenic for the male-specific T cell receptor (TCR) HY (HY/gammac- mice). In HY/gammac- male mice, negative selection of autoreactive thymocytes was not diminished; however, peripheral T cells expressing transgenic TCR-alpha and -beta chains (TCR-alphaT/betaT) were absent, and extrathymic T cell development was completely abrogated. In HY/gammac- female mice, the expression of the transgenic TCR partially reversed the profound thymic hypoplasia observed in nontransgenic gammac- mice, generating increased numbers of thymocytes in all subsets, particularly the TCR-alphaT/betaT CD8+ single-positive thymocytes. Despite efficient positive selection, however, naive CD8+ TCR-alphaT/betaT T cells were severely reduced in the peripheral lymphoid organs of HY/gammac- female mice. These results not only underscore the indispensible role of gammac in thymocyte development, but also demonstrate the critical role of gammac in the maintenance and/or expansion of peripheral T cell pools.

Two gut intraepithelial CD8+ lymphocyte populations with different T cell receptors: a role for the gut epithelium in T cell differentiation.

Mouse gut intraepithelial lymphocytes (IEL) consist mainly (90%) of two populations of CD8+ T cells. One bears heterodimeric alpha/beta CD8 chains (Lyt-2+, Lyt-3+), a T cell receptor (TCR) made of alpha/beta chains, and is Thy-1+; it represents the progeny of T blasts elicited in Peyer's patches by antigenic stimulation. The other bears homodimeric alpha/alpha CD8+ chains, contains no beta chain mRNA, and is mostly Thy-1- and TCR-gamma/delta + or -alpha/beta +; it is thymo-independent and does not require antigenic stimulation, as shown by its presence: (a) in nude and scid mice; (b) in irradiated and thymectomized mice repopulated by T-depleted bone marrow cells bearing an identifiable marker; (c) in thymectomized mice treated by injections of monoclonal anti-CD8 antibody, which lead to total depletion of peripheral CD8+ T lymphocytes; and (d) in germ-free mice and in suckling mice. In young nude mice, alpha/alpha CD8 chains, CD3-TCR complexes, and TCR mRNAs (first gamma/delta) are found on IEL, while they are not detectable on or in peripheral or circulating lymphocytes or bone marrow cells. IEL, in contrast to mature T cells, contain mRNA for the RAG protein, which is required for the rearrangement of TCR and Ig genes. We propose that the gut epithelium (an endoderm derivative, as the thymic epithelium) has an inductive property, attracting progenitors of bone marrow origin, and triggering their TCR rearrangement and alpha/alpha CD8 chains expression, thus giving rise to a T cell population that appears to belong to the same lineage as gamma/delta thymocytes and to recognize an antigenic repertoire different from that of alpha/beta CD8+ IEL.

Different use of T cell receptor transducing modules in two populations of gut intraepithelial lymphocytes are related to distinct pathways of T cell differentiation.

Most gut intraepithelial cells (IEL) of the mouse are T cells that bear CD8 molecules, present either as alpha-beta chain heterodimers (CD8 beta+) or as alpha chain homodimers (CD8 beta-). All CD8 beta+ IEL bear alpha/beta T cell receptors (TCR); CD8 beta- IEL bear either alpha/beta or gamma/delta TCR and are considered to be a thymus-independent (TI) population, probably arising locally from a small fraction of CD3- IEL containing the recombinant activating gene RAG proteins. Here we report that TI CD8 beta- IEL, whether bearing alpha/beta or gamma/delta TCR, contain, in normal mice, mRNAs for both zeta and Fc epsilon RI gamma chains. These chains are present in their CD3-TCR complexes as homo- or heterodimers. In contrast, only zeta chain mRNA and homodimers are found in gut CD8 alpha/beta+ IEL and in peripheral T lymphocytes. Intestinal CD3- precursor cells contain only gamma chain, and CD3- IL-2R+ thymocyte precursors only zeta chain mRNAs. Only very primitive thymocyte precursors contain detectable gamma chain mRNA, and it thus appears that Fc epsilon RI gamma chain use is switched off at a very early stage during thymocyte differentiation. Thus, T cell differentiation in the gut epithelium differs from that occurring in the thymus, from which CD8 beta+ IEL appear to derive. Use of different TCR transducing modules and CD8 accessory molecules between the TI and the thymus-derived T cell populations provides an explanation for their difference in reactivity to antigenic stimulations and thus in selection of repertoires.

Identical T cell clones are located within the mouse gut epithelium and lamina propia and circulate in the thoracic duct lymph.

Murine gut intraepithelial (IEL) T cell receptor (TCR)-alpha/beta lymphocytes bearing CD8alpha/13 or CD8alpha/alpha coreceptors have been shown previously to express different oligoclonal TCR beta chain repertoires in the same mouse, in agreement with other evidence indicating that these two populations belong to different ontogenic lineages, with only CD8alpha/beta+ IELs being fully thymus dependent. CD8alpha/beta+, but not CD8alpha/alpha+, T lymphocytes are also present in the lamina propria. Here, we show that CD8alpha/beta+ lymphocytes from the lamina propria and the epithelium are both oligoclonal, and that they share the same TCR-beta clonotypes in the same mouse, as is also the case for CD4alpha T cells. Furthermore, identical T cell clones were detected among CD8alpha/beta IELs and CD8alpha/beta+ blasts circulating into the thoracic duct (TD) lymph of the same mouse, whereas TD small lymphocytes are polyclonal. These findings must be considered in light of previous observations showing that T blasts, but not small T lymphocytes, circulating in the TD lymph have the capacity of homing into the gut epithelium and lamina propria. These combined observations have interesting implications for our understanding of the recirculation of gut thymus-dependent lymphocytes and their precursors, and of the events leading up to the selection of their restricted TCR repertoire.

Selection and expansion of CD8alpha/alpha(1) T cell receptor alpha/beta(1) intestinal intraepithelial lymphocytes in the absence of both classical major histocompatibility complex class I and nonclassical CD1 molecules.

Intestinal intraepithelial lymphocytes (IELs) in mice include two main subsets of TCR-alpha/beta(1) cells which differ functionally and ontogenically from each other. One expresses the CD8alpha/alpha homodimer, whereas the other expresses the CD8alpha/beta heterodimer. Although the presence of all CD8(+)TCR-alpha/beta(1) IELs is dependent on beta2-microglobulin molecules, the nature of the major histocompatibility complex (MHC) class I molecules recognized by the CD8alpha/alpha and the CD8alpha/beta(1) subsets has remained elusive. Using mutant mice lacking the expression of both H2-K(b) and H2-D(b), we show that the CD8alpha/beta(1)TCR-alpha/beta(1) subset is dependent on K or D molecules, whereas the CD8alpha/alpha(1)TCR-alpha/beta(1) subset is independent of classical MHC class I molecules. Furthermore, the CD8alpha/alpha(1) cells are conserved in mice lacking expression of CD1, a nonclassical MHC class I-like molecule previously proposed to be a potential ligand for IELs. Using transporter associated with antigen processing (TAP)-deficient mice, this cell population can be further separated into a TAP-dependent and a TAP-independent subset, suggesting either the recognition of two nonclassical MHC-like molecules, only one of which is TAP dependent, or the involvement of a single nonclassical MHC-like molecule that is only partially TAP dependent. These findings demonstrate that CD8alpha/beta(1)TCR-alpha/beta(1) IELs are restricted by H-2K and H-2D molecules, whereas the unusual subset of CD8alpha/alpha(1)TCR-alpha/beta(1) resident IELs recognize nonclassical MHC class I-like molecules that are distinct from CD1.

Contrast-to-noise ratios of liver lesions using subtraction imaging on multiphase 64-detector row CT.

AIM: To measure contrast-to-noise ratios of liver lesions on conventional enhanced and digitally subtracted multidetector row computed tomography (CT) images. MATERIALS/METHODS: This study was approved by our hospital internal review board (IRB) and all collected data were evaluated in a Health Insurance Portability and Accountability Act (HIPAA)-compliant manner. Subtracted datasets, using pixel-by-pixel subtraction of the post-contrast images from the pre-contrast images, were created from the 64 detector-row CT of patients undergoing three-phase examination of the liver (unenhanced, arterial phase, and portal venous phase). Regions of interest were used to calculate the contrast-to-noise ratios between the lesions and the background liver parenchyma on both the post-contrast and subtracted datasets using the following formula: (Lesion mean (HU) - Liver mean (HU))/standard deviation of mean outside patient (HU). These ratios were compared using a mixed linear statistical model. RESULTS: Contrast-to-noise ratios were calculated for 64 lesions in 50 consecutive patients. Of the 64 lesions, 42 were hypervascular and 22 were hypovascular. Subtracted datasets yielded statistically significant higher contrast-to-noise ratios of hypervascular lesions compared to normal liver parenchyma (p<0.0001). Subtraction did not yield a statistically significant improvement in contrast-to-noise ratios for hypovascular liver lesions (p=0.16). CONCLUSION: Post-processed subtraction CT images generate increased contrast-to-noise ratios for hypervascular liver lesions. As this technique is easy to perform and does not involve additional radiation exposure, it should be considered when evaluating for suspected hypervascular lesions.