An atlas of B-cell clonal distribution in the human body.

B-cell responses result in clonal expansion, and can occur in a variety of tissues. To define how B-cell clones are distributed in the body, we sequenced 933,427 B-cell clonal lineages and mapped them to eight different anatomic compartments in six human organ donors. We show that large B-cell clones partition into two broad networks-one spans the blood, bone marrow, spleen and lung, while the other is restricted to tissues within the gastrointestinal (GI) tract (jejunum, ileum and colon). Notably, GI tract clones display extensive sharing of sequence variants among different portions of the tract and have higher frequencies of somatic hypermutation, suggesting extensive and serial rounds of clonal expansion and selection. Our findings provide an anatomic atlas of B-cell clonal lineages, their properties and tissue connections. This resource serves as a foundation for studies of tissue-based immunity, including vaccine responses, infections, autoimmunity and cancer.

Dendritic Cells Display Subset and Tissue-Specific Maturation Dynamics over Human Life.

Maturation and migration to lymph nodes (LNs) constitutes a central paradigm in conventional dendritic cell (cDC) biology but remains poorly defined in humans. Using our organ donor tissue resource, we analyzed cDC subset distribution, maturation, and migration in mucosal tissues (lungs, intestines), associated lymph nodes (LNs), and other lymphoid sites from 78 individuals ranging from less than 1 year to 93 years of age. The distribution of cDC1 (CD141hiCD13hi) and cDC2 (Sirp-α+CD1c+) subsets was a function of tissue site and was conserved between donors. We identified cDC2 as the major mature (HLA-DRhi) subset in LNs with the highest frequency in lung-draining LNs. Mature cDC2 in mucosal-draining LNs expressed tissue-specific markers derived from the paired mucosal site, reflecting their tissue-migratory origin. These distribution and maturation patterns were largely maintained throughout life, with site-specific variations. Our findings provide evidence for localized DC tissue surveillance and reveal a lifelong division of labor between DC subsets, with cDC2 functioning as guardians of the mucosa.

Tissue reservoirs of antiviral T cell immunity in persistent human CMV infection.

T cell responses to viruses are initiated and maintained in tissue sites; however, knowledge of human antiviral T cells is largely derived from blood. Cytomegalovirus (CMV) persists in most humans, requires T cell immunity to control, yet tissue immune responses remain undefined. Here, we investigated human CMV-specific T cells, virus persistence and CMV-associated T cell homeostasis in blood, lymphoid, mucosal and secretory tissues of 44 CMV seropositive and 28 seronegative donors. CMV-specific T cells were maintained in distinct distribution patterns, highest in blood, bone marrow (BM), or lymph nodes (LN), with the frequency and function in blood distinct from tissues. CMV genomes were detected predominantly in lung and also in spleen, BM, blood and LN. High frequencies of activated CMV-specific T cells were found in blood and BM samples with low virus detection, whereas in lung, CMV-specific T cells were present along with detectable virus. In LNs, CMV-specific T cells exhibited quiescent phenotypes independent of virus. Overall, T cell differentiation was enhanced in sites of viral persistence with age. Together, our results suggest tissue T cell reservoirs for CMV control shaped by both viral and tissue-intrinsic factors, with global effects on homeostasis of tissue T cells over the lifespan.

Early-life compartmentalization of human T cell differentiation and regulatory function in mucosal and lymphoid tissues.

It is unclear how the immune response in early life becomes appropriately stimulated to provide protection while also avoiding excessive activation as a result of diverse new antigens. T cells are integral to adaptive immunity; mouse studies indicate that tissue localization of T cell subsets is important for both protective immunity and immunoregulation. In humans, however, the early development and function of T cells in tissues remain unexplored. We present here an analysis of lymphoid and mucosal tissue T cells derived from pediatric organ donors in the first two years of life, as compared to adult organ donors, revealing early compartmentalization of T cell differentiation and regulation. Whereas adult tissues contain a predominance of memory T cells, in pediatric blood and tissues the main subset consists of naive recent thymic emigrants, with effector memory T cells (T(EM)) found only in the lungs and small intestine. Additionally, regulatory T (T(reg)) cells comprise a high proportion (30-40%) of CD4(+) T cells in pediatric tissues but are present at much lower frequencies (1-10%) in adult tissues. Pediatric tissue T(reg) cells suppress endogenous T cell activation, and early T cell functionality is confined to the mucosal sites that have the lowest T(reg):T(EM) cell ratios, which suggests control in situ of immune responses in early life.

[Surgical treatment of acute cholecystitis].

Fundamental treatment for acute cholecystitis is cholecystectomy. However, the adoption of a treatment is dependend on degree of a severity of acute cholecystitis in each patient because its degree is influenced by factors such as duration from the onset of symptoms to medical examination. Early laparoscopic cholecystectomy is the preferred procedure for mild acute cholecystitis. Early cholecystectomy is also performed for moderately acute cholecystitis. However, if patients have severe local inflammation (gangrenous and purulent cholecystitis) early gallbladder drainage or open cholecystectomy is indicated. Emergency operation under adequate medical treatment is indicated for a patient with severe local inflammation of the gallbladder, torsion of the gallbladder, emphysematous cholecystitis, gangrenous cholecystitis, and purulent cholecystitis. Pericholecystic abscess, necrosis of the gallbladder wall, and perforation of the gallbladder can be diagnosed accurately by use of imaging diagnosis. The optimal surgical treatment for acute cholecystitis according to grade of severity should be performed referring to imaging findings.

Prevention of high-mobility group box 1-mediated early loss of transplanted mouse islets in the liver by antithrombin III.

BACKGROUND: The low efficiency of pancreatic islet transplantation mainly because of the early loss of transplanted islets hampers its clinical application. Previously, we have shown in mice that the early loss of transplanted islets in the liver is caused by innate immune rejection in concert with dendritic cells, natural killer T cells, and neutrophils to produce interferon (IFN)-γ, which is triggered by high-mobility group box 1 (HMGB1) released from transplanted islets. We herein determined whether the HMGB1-mediated early loss of transplanted mouse islets is prevented by antithrombin (ATIII). METHODS: The effect of ATIII on in vitro and in vivo HMGB1-stimulated IFN-γ production of hepatic mononuclear cells was examined. Then, the effect of ATIII on amelioration of hyperglycemia in streptozotocin-induced diabetic mice receiving 200 syngeneic islets from a single donor was determined. RESULTS: In vitro and in vivo IFN-γ production of mononuclear cells in the liver of mice in response to HMGB1 was suppressed by ATIII. Hyperglycemia of streptozotocin-induced diabetic mice receiving 200 syngeneic islets into the liver from a single donor was ameliorated with down-regulation of IFN-γ production of natural killer T cells and neutrophils in the liver when ATIII but not vehicle was administered once at the time of islet transplantation. The favorable effect of ATIII was similarly achieved in mice receiving islet allografts when rejection was prevented with anti-CD4 antibody treatment. CONCLUSIONS: These findings demonstrate that ATIII prevents HMGB1-mediated early loss of transplanted islets caused by innate immune rejection, suggesting a potential application of ATIII to improve efficiency of clinical islet transplantation.

Single orifice vein reconstruction in left liver plus caudate lobe graft.

BACKGROUND/AIMS: We describe the innovative techniques with single orifice vein reconstruction for the complete venous drainage in the left liver plus caudate lobe graft. METHODOLOGY: Eight left liver plus caudate lobe grafts used for living donor adult liver transplantation were reviewed. A wide and single venous orifice was created by gathering the left, middle or its tributaries, and/or short hepatic vein using a patch vein graft or a conduit vein graft. This single, newly-created orifice was then anastomosed to the common trunk created in the recipient's hepatic veins. RESULTS: Of 8 liver grafts, six included the middle hepatic vein trunk. Another two included only the middle hepatic vein tributaries. Significantly-sized short hepatic veins were preserved in 4 grafts and were connected with the major hepatic veins or tributaries of the middle hepatic vein to make a single orifice using a conduit vein graft. For remaining 4 grafts without significantly-sized short hepatic veins, two adjacent hepatic vein trunks (left and middle hepatic veins) were simply connected together. To enlarge the common orifice of the hepatic veins, the patch vein grafts were further attached. Hepatic vein waveforms of all grafts showed the biphasic or triphasic pattern and the graft congestion was not observed immediately after venoplasty. No graft was lost due to hepatic venous outflow block with the mean follow-up of 15 months. CONCLUSIONS: The short-term results of our technique were satisfactory. The present technique can simplify graft-to-recipient hepatic vein reconstruction without unfavorable tension on the anastomosis.

Limy bile syndrome complicated by obstructive jaundice.

Limy bile syndrome is a rare condition in which the gallbladder is filled with a paste-like radiopaque material. The presence of limy bile in the common bile duct is rare. A 72-year-old woman was admitted to our hospital with epigastric pain and jaundice. Plain abdominal radiography on admission showed a radiopaque material in the gallbladder. Computed tomography also showed that the gallbladder and the common bile duct were filled with a radiopaque material. The patient had never received any cholecystographic contrast agents. As a result, a diagnosis of obstructive jaundice due to choledocholithiasis, which includes limy bile, was made. We herein report the process by which limy bile syndrome, complicated by obstructive jaundice, was successfully treated through combined treatment via endoscopic sphincterotomy and laparoscopic cholecystectomy.

Impact of short hepatic vein reconstruction in living donor adult liver transplantation using a left liver plus caudate lobe graft.

OBJECTIVE: To investigate the impact of short hepatic vein reconstruction in the transplanted left liver plus caudate lobe graft. METHODS: Six left liver plus caudate lobe grafts used for living donor adult liver transplantation were included in this study. The liver grafts were divided into two groups: those with (V1 group; n = 4) or without (control group; n = 2) short hepatic vein reconstruction. The changes in the transplanted left lobe (segments II-IV) and caudate lobe were compared between the two groups at 1 month after transplantation. RESULTS: The addition of the caudate lobe increased the graft volume by 15 mL, which corresponded to a 4.3% gain of graft volume at the time of transplantation. Although the graft volume/standard liver volume ratio of the whole grafts after transplantation showed no difference between the two groups, the regeneration rate of the caudate lobe in the V1 group was significantly greater than that in the control group (p= 0.04). CONCLUSION: Although no definite advantage from the V1 reconstruction was demonstrated, hepatic vein reconstruction with a significantly-sized short hepatic vein might provide an additional margin of safety for marginally-sized liver grafts during the early phase of graft regeneration.

High-mobility group box 1 is involved in the initial events of early loss of transplanted islets in mice.

Islet transplantation for the treatment of type 1 diabetes mellitus is limited in its clinical application mainly due to early loss of the transplanted islets, resulting in low transplantation efficiency. NKT cell-dependent IFN-gamma production by Gr-1(+)CD11b(+) cells is essential for this loss, but the upstream events in the process remain undetermined. Here, we have demonstrated that high-mobility group box 1 (HMGB1) plays a crucial role in the initial events of early loss of transplanted islets in a mouse model of diabetes. Pancreatic islets contained abundant HMGB1, which was released into the circulation soon after islet transplantation into the liver. Treatment with an HMGB1-specific antibody prevented the early islet graft loss and inhibited IFN-gamma production by NKT cells and Gr-1(+)CD11b(+) cells. Moreover, mice lacking either of the known HMGB1 receptors TLR2 or receptor for advanced glycation end products (RAGE), but not the known HMGB1 receptor TLR4, failed to exhibit early islet graft loss. Mechanistically, HMGB1 stimulated hepatic mononuclear cells (MNCs) in vivo and in vitro; in particular, it upregulated CD40 expression and enhanced IL-12 production by DCs, leading to NKT cell activation and subsequent NKT cell-dependent augmented IFN-gamma production by Gr-1(+)CD11b(+) cells. Thus, treatment with either IL-12- or CD40L-specific antibody prevented the early islet graft loss. These findings indicate that the HMGB1-mediated pathway eliciting early islet loss is a potential target for intervention to improve the efficiency of islet transplantation.

Prevention of early loss of transplanted islets in the liver of mice by adenosine.

BACKGROUND: The low efficiency of islet transplantation necessitating sequential transplantations with the use of 2 to 3 donors for a recipient has been a major obstacle facing clinical islet transplantation. We determined whether adenosine has any beneficial effects on preventing early loss of transplanted islets in the liver, thereby facilitating successful islet transplantation from one donor to one recipient in mice. METHODS: Two hundred islets, the number of islets from a single mouse pancreas, were grafted into the liver of streptozotocin-induced diabetic C57BL/6 mice. Adenosine was administered once at the time of islet transplantation. Mononuclear cells in the liver of mice receiving islets were isolated and examined by flow cytometry. RESULTS: A single injection of adenosine at the time of transplantation ameliorated hyperglycemia of diabetic mice receiving 200 syngenic islets with suppression of interferon (IFN)-gamma production of hepatic NKT cells and neutrophils, while that of control did not. The IFN-gamma production of NKT cells and neutrophils in the liver of mice treated with alpha-galactosylceramide, a synthetic ligand of NKT cells was suppressed by adenosine. The beneficial effect of adenosine was also observed for BALB/c islet allografts when alloimmune rejection was prevented by anti-CD4 antibody. CONCLUSIONS: Adenosine suppresses the NKT cell-mediated IFN-gamma production of neutrophils in the liver of mice receiving islets, thus leading to prevention of early loss of transplanted syngenic and allogenic islets. The findings indicate that adenosine may improve efficiency of clinical islet transplantation.

Successful islet transplantation to two recipients from a single donor by targeting proinflammatory cytokines in mice.

BACKGROUND: Currently, the inability to achieve successful islet transplantation from one donor to one recipient is a major obstacle facing clinical islet transplantation. We herein determined whether this limitation could be overcome by targeting pro-inflammatory cytokines with the prevention of immediate islet graft loss in association with engraftment in mice. METHODS: Isolated islets were grafted into the liver of streptozotocin-induced diabetic mice and the role of proinflammatory cytokines in the engraftment of islets was evaluated with the use of interferon (IFN)-gamma-/- mice and monoclonal antibodies against proinflammatory cytokines. RESULTS: Hyperglycemia in streptozotocin-induced diabetic mice receiving 200 syngenic islets, which were isolated from a single mouse pancreas, was ameliorated when IFN-gamma-/-, but not wild-type mice, were used as recipients. The treatment with anti-IFN-gamma antibody produced normoglycemia in diabetic wild-type mice receiving 200, but not 100 islets. However, when anti-tumor necrosis factor-alpha and anti-interleukin-1beta antibodies were administered in conjunction with anti-IFN-gamma antibody, wild-type diabetic mice receiving 100 islets became normoglycemic after transplantation. In addition, the favorable effect of the combined use of antibodies was similarly achieved in mice receiving islet allografts when rejection was prevented with anti-CD4 antibody treatment. CONCLUSIONS: These findings clearly demonstrate that successful islet transplantation from one donor to two recipients is feasible by targeting pro-inflammatory cytokines in mice, thus suggesting a potential application in clinical islet transplantation if similar mechanisms of islet graft loss could be mediated in humans.

Natural killer T-cells participate in rejection of islet allografts in the liver of mice.

A role of natural killer T (NKT) cells in transplant rejection remains unknown. Here, we determined whether NKT cells participate in rejection of islet allografts, using NKT cell-deficient mice. Survival of islet allografts in streptozotocin-induced diabetic CD1d(-/-) mice or Valpha14 NKT cell(-/-) mice was significantly prolonged without immunosuppression when grafted into the liver, but not beneath the kidney capsule, compared with wild-type mice. Acceptance of intrahepatic islet allografts was achieved in CD1d(-/-) mice by a subtherapeutic dose of rapamycin, which was abrogated in conjunction with the transfer of hepatic mononuclear cells from wild-type, but not from CD1d(-/-), mice at islet transplantation. The second islet grafts from a donor-specific, but not from a third-party, strain in CD1d(-/-) mice bearing functional islet allografts were accepted without immunosuppression at 120 days after the initial transplantation. These findings demonstrate that NKT cells play a significant role in rejection of islet allografts in the liver of mice, but that NKT cells are not essential for induction of donor-specific unresponsiveness in this model. The current study indicates that NKT cells might be considered as a target for intervention to prevent islet allograft rejection when the liver is the site of transplantation.

Valpha14 NK T cell-triggered IFN-gamma production by Gr-1+CD11b+ cells mediates early graft loss of syngeneic transplanted islets.

Pancreatic islet transplantation is a highly promising approach for the treatment of insulin-dependent diabetes mellitus. However, the procedure remains experimental for several reasons, including its low efficiency caused by the early graft loss of transplanted islets. We demonstrate that Gr-1+CD11b+ cells generated by transplantation and their IFN-gamma production triggered by Valpha14 NKT cells are an essential component and a major cause of early graft loss of pancreatic islet transplants. Gr-1+CD11b+ cells from Valpha14 NKT cell-deficient (Jalpha281-/-) mice failed to produce IFN-gamma, resulting in efficient islet graft acceptance. Early graft loss was successfully prevented through the repeated administration of alpha-galactosylceramide, a specific ligand for Valpha14 NKT cells, resulting in dramatically reduced IFN-gamma production by Gr-1+CD11b+ cells, as well as Valpha14 NKT cells. Our study elucidates, for the first time, the crucial role of Gr-1+CD11b+ cells and the IFN-gamma they produce in islet graft rejection and suggests a novel approach to improving transplantation efficiency through the modulation of Valpha14 NKT cell function.

Beneficial effects of costimulatory blockade with anti-inducible costimulator antibody in conjunction with CTLA4Ig on prevention of islet xenograft rejection from rat to mouse.

BACKGROUND: Costimulatory signals have been reported to play an important role in islet-xenograft rejection, although the precise mechanisms remain unknown. The aim of the present study was to determine a role of a novel costimulatory molecule, inducible costimulator (ICOS), in rat islet-xenograft rejection in conjunction with CTLA4Ig with respect to cellular as well as humoral immune responses. METHODS: Isolated rat islets were transplanted into the liver of streptozotocin (180 mg/kg) induced diabetic mice. Cellular immune responses to islet xenografts, and productions of anti-rat antibody in mice were examined by flow cytometry (FACS) after transplantation. RESULTS: Intrahepatic rat islet xenografts were rejected in mice within 8 days after transplantation. FACS analysis revealed an expansion of CD8(+) T cells in the liver as well as a production of anti-rat antibody in recipient mice in association with rejection. The treatment with anti-ICOS antibody in conjunction with CTLA4Ig produced a marked prolongation of islet-xenograft survival with neither expansion of CD8(+) T cells nor production of anti-rat antibody, whereas, in contrast, those treated with anti-ICOS antibody or CTLA4Ig alone did not have prolonged survival, and CD8(+) T cells were expanded. CONCLUSION: These findings demonstrate that cellular rather than humoral immune responses are considered responsible for islet-xenograft rejection from rat to mouse and that the blockade of costimulatory signals with anti-ICOS antibody in conjunction with CTLA4Ig has a favorable effect on prevention of islet xenograft rejection.