池住 洋平

わが国では小児IgA腎症の治療指針が示され、比較的重症例に対してステロイド薬、免疫抑制薬を含むカクテル療法が行われ良好な成果が得られている。しかし、治療による副作用などマイナス面については十分に検討されていない。今回私たちは、ステロイド治療が成長に与える影響を後方視的に検討した。対象はステロイド治療を行った小児IgA腎症48例(治療開始時平均11歳10ヵ月)で、2年間の治療前後で身長、体重および骨年齢を比較した。身長SDスコアは平均0.229から-0.116(p<0.05)へと有意に低下した。一方、肥満度は平均-0.38から5.94(p<0.05)へと有意な増加を認め、骨塩量SDスコアは平均-0.357から-0.853と有意差はないものの低下傾向を認めた。小児IgA腎症のステロイド治療により成長障害を高頻度に来す可能性があり、今後はこうした副作用を考慮した治療法の質的向上が必要と考えられた。(著者抄録)

Hypotonic fluids are commonly used for treating hospitalized children. However, an excess of arginine vasopressin (AVP) with impaired free water excretion is thought to contribute to the development of hyponatremia in febrile children. The aim of this two-part study was to define the clinical relationship between hyponatremia and excess AVP. In a retrospective study carried out between 2001 and 2005, we found that approximately 17% of the hospitalized patients had hyponatremia [serum sodium (Na) < 135 mEq/l] upon admission and that the ratio of patients with hyponatremia was significantly higher among febrile patients than among afebrile patients. In a subsequent prospective study, we examined 73 hospitalized patients who presented with acute febrile diseases accompanied by hyponatremia (serum Na < 134 mEq/l). Almost all of these patients demonstrated excess AVP, defined as high plasma AVP levels (> 1 pg/ml). There were no significant relationships between the levels of AVP and other laboratory variables, including serum sodium, serum osmolality, atrial natriuretic peptide, and brain natriuretic peptide. About 30% (22/73) of the patients fulfilled the criteria of the syndrome of inappropriate secretion of antidiuretic hormone. These findings suggest that fever and other nonosmotic stimuli lead directly to excess AVP and hyponatremia. We therefore recommend that isotonic fluids should be used for patients with prolonged fever and hyponatremia.

今後の小児chronic kidney disease(CKD)に対する薬物療法のエビデンス作りを目的に、成人の糖尿病性腎症以外のCKDに対するangiotensin-converting enzyme inhibitor(ACEI)とangiotensin receptor blocker(ARB)の腎保護作用にについて文献的考察を行った。その結果、Medlineと医学中央雑誌の過去10年間の文献検索により、成人のCKDに対するACEIとARBの効果に関する文献が多数検出され、その多くは有効性を報告する結論であった。ACEIとARBの比較では同等又はARBの方が有効とする報告が散見され、ACEIとARBの単独療法と併用療法の比較では、併用療法の方がより腎保護効果が高いとする報告もみられた。このように成人のCKDに対するACEIとARBの有効性がエビデンスを持って多数報告されていたが、小児のCKDに対する有効性についてのエビデンスは認められなかった。以上より、小児のCKDに対するエビデンスの蓄積が急務であり、今後適切にデザインされたランダム化比較試験の実施が必要と考えられた。

The development of proteinuria and glomerulosclerosis in kidney disease is associated with podocyte damage, including down-regulation of nephrin and podocin. Macrophages are known to induce renal injury, but the mechanisms involved are not fully understood. This study examined macrophage-mediated podocyte damage. Conditioned media (CM) from activated macrophages caused a 50-60% reduction in nephrin and podocin nnRNA and protein expression in cultured mouse podocytes and rat glomeruli. This was abolished by a neutralizing anti-TNF alpha antibody. The addition of recombinant TNF alpha to podocytes OF glomeruli caused a comparable reduction in podocyte nephrin and podocin expression to that of macrophage CM. Inhibition of c-Jun amino terminal kinase (JNK) or p38 kinase abolished the TNF alpha-induced reduction in nephrin and podocin expression. This study demonstrates that activated macrophages can induce podocyte injury via a TNF alpha-JNK/p38-dependent mechanism. This may explain, in part, the protective effects of JNK and p38 blockade in experimental kidney disease. (C) 2008 Elsevier Inc. All rights reserved.

わが国では法制度のもとに学校検尿が実施され、小児腎疾患の早期発見と治療成績の向上に貢献してきた。しかし、本制度に基づく疫学調査がほとんど行われていないのが現状である。また、小児腎疾患として頻度の高い特発性ネフローゼ症候群についても、全国規模の疫学データがほとんどない。今回私たちは、新潟市における学校検尿に基づく小児IgA腎症の疫学調査および新潟県内ネットワークを利用した小児ネフローゼ症候群の疫学調査を試みた。1993年から2006年の新潟市内の学校検尿受診者を対象に調査を行ったところ、受検者10,000人あたり毎年約0.68人の新規発症IgA腎症患者が学校検尿で発見されていることが明らかとなった。また、学校検尿で発見される症例(学校検尿群)は肉眼的血尿等で直接医療機関に受診した患者(肉眼的血尿群)より、発症から腎生検までの観察期間が有意に長く、治療2年後の寛解率は有意に高かった。新潟県内のネフローゼ症候群は毎年15歳未満の小児10万人あたり平均4.2人が新規に発症していた。この頻度は約40年前とほぼ同等であったが、小児人口の減少によって、有病数は約3分の2に大きく減少していると推測された。学校検尿制度を実施しているわが国は、他国では不可能なデータ解析を可能とし、疫学調査のみならず臨床上の有益な情報収集に寄与するものと考えられた。また、情報ネットワークの構築とその拡大が全国規模の疫学調査には不可欠であると考えられた。(著者抄録)

ABO不適合腎移植は、一般的に脾臓摘出が必須と考えられてきたが、小児においてはデメリットが大きい。今回われわれは、ABO不適合腎移植を希望した12歳男児に対し、移植4週間前からMMF,MPによる脱感作療法を開始し、12日前と2日前にrituximabの投与を加え脾臓摘出の回避を試みた。重篤な副作用はなく、血液型抗体の再上昇を認めず、移植後3年の時点で経過は良好である。同じプロトコルで行った成人例に比べるとB細胞の回復は早い傾向にあったが、免疫学的順応が得られる数週間の抑制を目的としている観点からは、まだ過剰抑制の可能性がある。今後は小児における必要最低限のRituximab投与方法の検討が必要と思われる。(著者抄録)

Aim: A number of growth factors have been shown to induce proliferation of renal cell types in animal models of kidney disease. In vitro studies suggest that many such growth factors induce renal cell proliferation through the extracellular signal-regulated kinase (ERK) pathway. The aim of this study was to determine the functional role of ERK signalling in cell proliferation in the obstructed kidney. Methods: Unilateral ureteric obstruction was induced in C57BL/6J mice which then received an ERK inhibitor drug (U0126 100 mg/kg t.i.d.), vehicle (DMSO) or no treatment, starting at day 2 after unilateral ureteric obstruction surgery and continuing until animals were killed on day 5. Cell proliferation was assessed by uptake of bromodeoxyuridine (BrdU). Results: In normal mice, phosphorylation (activation) of ERK (p-ERK) was restricted to collecting ducts. Western blotting identified a marked increase in p-ERK in the obstructed kidney in the no-treatment and vehicle-treated groups. Immunostaining showed strong p-ERK staining in many tubules and in interstitial cells. U0126 treatment inhibited ERK phosphorylation as assessed by western blot and immunostaining. The number of BrdU+ cortical tubular cells was reduced by vehicle treatment but was not further changed by U0126 treatment. In contrast, interstitial cell proliferation in the obstructed kidney was unaltered by vehicle treatment, but this was significantly inhibited by U0126. This was associated with a reduction in interstitial macrophage accumulation, but no effect was seen upon interstitial accumulation of alpha-SMA+ myofibroblasts. Renal fibrosis, as assessed by collagen deposition, was unaffected by U0126 or vehicle treatment. Conclusion: These studies show that accumulation of interstitial macrophages in the obstructed kidney is, in part, dependent upon the ERK signalling pathway.

2006年までの10年間に報告された成人のCKDに対するACEIとARBの腎保護作用に関する文献47編(英文38編、和文9編)と小児の場合の腎保護作用および高血圧に対する治療効果に関する文献18編をMedlineおよび医学中央雑誌より検索してレビューを行った。成人に関する英文3編はmeta analysisで29編(英文25、和文4)はランダム化比較試験(RCT)文献であった。小児の腎保護作用に関する文献はなく、RCT文献4編は全て高血圧に対する治療効果の検討であり、CKDに対する腎保護作用に関するRCTはなかった。成人の非糖尿病性腎症に対するACEIの腎保護作用に関する文献20編はmeta analysis 3編、RCT 12編で、尿蛋白減少効果を検討した10編の文献はいずれも有効結果であった。成人のCKDに対するACEIやARBの併用療法の有効性の報告は多数検索され、ACEIとARBの比較では同等またはARBの方を有効とする報告が散見し、ACEIまたはARB単独よりACEI+ARB併用の腎保護効果が高いとする報告があった。成人の検討対象に多く使用された薬剤はACEIではenalapril、ramipril、lisinoprilなどでARBはlosartan、candesartan、valsartanで、糖尿病性腎症以外のCKDに対してもACEIとARBの有効性に関するエビデンス報告を認めた。小児の検討対象に使用された薬剤はACEIではenalapril、ramipril等でARBではlosartan、irbesartan等であった。小児のCKDに対するACEIやARBの有効性を示唆する報告は症例やコホート研究のみで、今後は小児の薬物療法に関するランダム化試験の実施が必要と考えられた。

Recent clinical trials have shown a beneficial effect of mizoribine (Miz), an immunosuppressive drug, in the treatment of new-onset pediatric IgA nephropathy (IgAN). In this study, we evaluated the efficacy of Miz treatment in three children with established steroid-resistant IgAN. The patients had IgAN featuring persistent proteinuria and diffuse mesangial proliferation and had failed to respond to 2 years of treatment with prednisolone. Based upon the second biopsy results, patients were given methylprednisolone (mPSL) pulse therapy that induced a transient reduction in proteinuria, which was reversed when the mPSL dose was tapered. Miz therapy was then instigated in place of pulse mPSL. All three patients showed a substantial reduction in proteinuria and resolution of hematuria within 5 months. A follow-up biopsy in two of the patients showed a substantial reduction in the severity of glomerular lesions and a decrease in the number of activated macrophages. In conclusion, Miz therapy was found to be a safe and effective therapy in three cases of steroid-resistant pediatric IgAN. The ability of Miz to reduce the number of activated macrophages may be an important mechanism by which this drug ameliorated renal disease in these patients.

EBV infection is one of major complications arising in pediatric patients who have undergone renal transplantation. A strong correlation between the grade of immunosuppression and the development of PTLD, one of the most severe EBV-associated diseases, has been recognized. In this study, we monitored the serologic profile in conjunction with peripheral blood EBV-DNA load of 32 children who underwent renal transplantation with tacrolimus as an immunosuppressant. Six patients were EBV-seronegative (EBV-) before the transplantation, and the mean DNA load in the EBV- group was significantly higher than that in the EBV-seropositive (EBV+) group. Seroconversion occurred in five of these patients in a mean period of 22 weeks after the transplantation. The EBV-DNA load in the EBV+ group was maintained at a low level for a year, whereas it increased rapidly to over 1 x 10(5) copies/mL in two patients in the EBV- group three to seven months after the transplantation, which corresponds to the timing of seroconversion, and one of them developed PTLD. These observations suggest that the close monitoring of the EBV-DNA load, along with longitudinal observation of seroconversion, is essential in pediatric renal transplantation, particularly for younger children who are more likely to be EVB-.

母および母方祖母が2型糖尿病という家族歴を有し、腎移植後糖尿病(PTDM)を発症した12歳、女児を経験した。術前のOGTTでは異常を認めず、タクロリムスベースの免疫抑制を開始していたが、術後1週間過ぎから高血糖となった。術後2ヵ月の時点でタクロリムスをシクロスポリンに変更した後の血糖コントロールは著明に改善し、インスリンから離脱することができた。少なくとも移植後早期のPTDMに対しては免疫抑制薬の変更が有効であると思われるとともに、本症例の経験から、1親等内に糖尿病の家族歴がある場合はシクロスポリンベースの免疫抑制が望ましいと考えた。(著者抄録)

Background. It is suggested that IgA nephropathy (IgAN) manifests differently in children vs adults on the basis of biopsy findings. However, this has been difficult to establish owing to the uncertainty of the timing of disease onset in adult IgAN. We addressed this question by comparing both histology and leucocyte accumulation in biopsies of recently diagnosed childhood and adult IgAN. Methods. Biopsies taken within 2 years from the onset of renal abnormalities in 33 childhood (10 +/- 3 years of age) and 38 adult (35 +/- 6 years) cases of IgAN were examined for histological changes (cellularity in mesangial, endocapillary and extracapillary areas, matrix expansion, adhesions/crescents and interstitial damage), glomerular deposition of immunoglobulin and complement, and the presence of macrophages, activated macrophages and T cells by immunohistochemistry. Results. Glomerular hypercellularity owing to increased cells in mesangial area was prominent in paediatric IgAN and significantly greater than in adult IgAN. In contrast, glomerular matrix expansion, crescent formation and interstitial damage were more severe in adults compared to paediatric IgAN. Indeed, glomerular hypercellularity correlated with proteinuria in paediatric but not in adult IgAN, whereas glomerular matrix correlated with proteinuria and renal function in adult but not in paediatric IgAN. The degree of C3c deposition was significantly greater in paediatric IgAN, while deposition of fibrinogen was greater in adult IgAN. Glomerular and interstitial CD68+ macrophages and a subset of sialoadhesin (Sn)+ activated macrophages were identified in both paediatric and adult IgAN, being significantly greater in number in adult IgAN. Glomerular leucocyte infiltration correlated with proteinuria while interstitial leucocyte infiltration correlated with interstitial damage in both groups. However, only the subset of Sn+ macrophages gave a significant correlation with renal function, glomerular hypercellularity and glomerular matrix. Conclusions. This study has demonstrated significant differences in the early glomerular lesions of IgAN in children vs adults. Furthermore, Sn+ activated macrophages are implicated in the pathogenesis of IgAN in both patient groups. The prognostic significance of these findings warrants further study.

Background. Sialoadhesin ( Sn; CD169) is a lectin-like receptor whose expression is restricted to subsets of tissue and inflammatory macrophages. We have previously identified accumulation of Sn+ macrophages as an important marker of disease progression versus remission in rat mesangial proliferative nephritis. The current study examined the significance of Sn+ macrophages in human proliferative glomerulonephritis. Methods. Frozen kidney sections from normal adult human kidney ( n = 4) and pediatric nephropathy ( n 40) were stained for total macrophages ( CD68+ cells), Sn+ macrophages, CD3+ T-cells and collagen type I by immunofluorescence. Leukocyte infiltration and the severity of glomerular lesions and interstitial damage were scored. A second protocol biopsy was performed in 27 cases and clinical and biopsy-based data obtained. Results. Sn+ macrophages were absent from glomeruli in normal adult human kidney and in thin basement membrane disease ( n = 4), but were detected in 4 of 9 cases of purpura nephritis; 7 of 17 IgA nephropathy; 5 of 5 membranoproliferative glomerulonephritis, and 5 of 5 lupus nephritis. Sn+ macrophages were localized in areas of focal glomerular and interstitial damage. Two-colour immunostaining confirmed that Sn+ cells are a subset of total CD68+ macrophages. The number of glomerular Sn+ macrophages correlated with the degree of proteinuria and glomerular lesions ( r = 0.44, P = 0.0045 and r = 0.82, P < 0.0001; respectively), while interstitial Sn+ macrophages correlated with the degree of proteinuria and interstitial damage ( r = 0.59, P < 0.0001 and r = 0.75, P < 0.0001; respectively). Combined immunostaining revealed that interstitial Sn+ macrophages and CD3+ T-cells co-localized in areas of tubulointerstitial damage with increased type I collagen deposition. There was significant correlation between the number of interstitial Sn+ macrophages and CD3+ T-cells ( r = 0.74, P < 0.0001). Most patients responded to a 2 year period of glucocorticoid therapy with a reduction in proteinuria and glomerular lesions and this correlated with the reduction in the number of glomerular Sn+ macrophages. Conclusion. This study has identified Sn+ cells as a macrophage subset whose accumulation in the kidney correlates with proteinuria and histologic damage. These results, together with recent findings from animal studies, suggest that Sn+ macrophages may play an important role in progressive renal disease.

経過中に体位性蛋白尿(OA)症状を呈し,かつ糸球体障害を有する16例(腎炎群)と合併所見のないOA14例(対照群)の尿中γグロブリン,FDP値を比較検討した.その結果,前彎負荷試験では両群とも10分間の負荷後に著明な蛋白尿の増加を認め,臥位安静で2時間までに消失または減少するOAパターンを示した.尿中γグロブリン値およびFDP値は,対照群ではいずれも著明な増加を認めたが,腎炎群ではいずれも有意に低値を示した.腎炎にみられるOA症状は,糸球体係蹄壁の構造異常が蛋白尿の発現機序に影響している可能性が示唆され,尿中蛋白分画およびFDP値の測定はOAの補助診断として有用と考えられた

20歳女.6歳時の学校検尿で血尿,蛋白尿を指摘され,ステロイド抵抗性のため腎生検を施行した.光顕ではメサンギウム領域を中心にPAS染色に淡染性の均一な無構造物質を認めた.電顕ではメサンギウム領域を中心に多量の線維の蓄積を認め,無秩序に配列し,特異形態を示していた.蛍光抗体法では一部IgM,C1q,C4が染まり,Collagenofibrotic glomerulopathyと診断した.治療抵抗性で高度の蛋白尿が持続し,腹膜透析を経て18歳時に父親をドナーとした生体腎移植を行った.明らかな拒絶反応はなく,血清クレアチニン値は0.7mg/dl前後で安定した.血中タイプIIIプロコラーゲン-N-ペプチド濃度により体内コラーゲン産生亢進を検討したところ,移植直後の1.2U/lから徐々に上昇し,7ヵ月後には4.0U/lとなり,その後は低下したが,術後12ヵ月以降も2.0U/l前後と高値で推移した.尿蛋白は認めず,移植腎機能は良好で,腎生検所見でも異常コラーゲンはみられなかった

Background. Glomerular accmulation of leukocytes, including lymphocytes, is a common feature in most types of glomerulonephritis. However, the role of lymphocytes in progressive glomerulonephritis has not been elucidated. We examined the role of lymphocytes in the development of progressive mesangial proliferative glomerulonephritis induced by two injections of monoclonal antibody 1-22-3 in rats. Methods. To elucidate the role of lymphocytes, circulating lymphocytes were depleted using specific monoclonal antibodies to rat lymphocytes prior to the induction of progressive glomerulonephritis. The effects of lymphocyte depletion on proteinuria and glomerular alterations were assessed 7 and 56 days after the induction of progressive glomerulonephritis. Results. Significant glomerular accmulation of CD4+ T cells, CD8+ T cells, and ED3+-activated macrophage were observed after the induction of glomerulonephritis. Depletion studies showed that continuous treatment with anti-CD5, anti-CD4, or anti-CD8 treatment reduced proteinuria and ameliorated the glomerular lesions on day 56. Depletion of CD4+ T cells also reduced glomerular accmulation of CD8+ T cells and ED3+-activated macrophages, and reduced glomerular expression of mRNA for interferon-gamma (INF-gamma) (63.0% in anti-CD5 and 62.3% reduction in anti-CD4). Transit lymphocyte depletion limited in early stage of progressive glomerulonephritis demonstrated that CD4+ T-cell depletion, but not anti-CD8 treatment prevented glomerular injuries 56 days after the induction of progressive glomerulonephritis. Conclusion. CD4+ T cells played a central role in the development of progressive glomerulonephritis, controlling recruitment and activation of CD8+ cytotoxic cells and/or macrophages.

Macrophage accumulation is a prominent feature in most forms of human glomerulonephritis and correlates with renal dysfunction. Macrophages can directly mediate acute renal injury in animal models, but the mechanisms of macrophage activation required for mediating renal injury are unknown. This study examined whether activation of the Jun amino terminal kinase (JNK) signaling pathway is necessary for macrophage-mediated renal injury. An adoptive transfer model was used in which rats were immunized with sheep IgG (day -5), made leukopenic by administration of cyclophosphamide (CyPh) (day -2), and then injected with sheep anti-glomerular basement membrane (GBM) serum (day 0). Animals were then given an intravenous injection of bone marrow-derived macrophages (BMM) (day 1) and killed 24 h later (day 2). The induction of proteinuria and glomerular cell proliferation (PCNA(+) cells) in CyPh-treated anti-GBM disease was dependent on transfer of BMM. Exposure of BMM to the specific JNK inhibitor, SP600125, for 3 h before adoptive transfer had no effect on glomerular accumulation of BMM in CyPh-treated anti-GBM disease. However, SP600125 treatment of BMM caused a 75% reduction in proteinuria and a 70% reduction in glomerular cell proliferation (P < 0.01 versus vehicle or untreated BMM). In conclusion, this study has defined a critical role for the JNK signaling pathway in macrophage-mediated renal injury.

19歳女.3歳時に精神発達遅滞と診断され,6歳時に学校検尿で血尿,蛋白尿を指摘されたことを契機にステロイド抵抗性であることから腎生検を施行し,collagenfibrotic glomerulopathyと診断した.各種治療に抵抗性を示し,13歳時に尿毒症症状が出現し腹膜透析を導入した.透析導入5年目でその効果が不十分となり,18歳時に父親をドナーとして生体腎移植を施行し,その後の経過は良好である.本症の症例学会発表を行った13施設を対象にアンケート調査を行い,9施設から回答を得た.診断時年齢は5〜65歳で15歳以下は本例を含め2例のみで,男女差はなく,平均観察期間7.2年で14例中7例が透析導入されていたが,腎移植例はなかった

Collagenofibrotic glomerulopathy is a recently recognized entity that is characterized by massive accumulation of collagen fibrils in the mesangial and subendothelial areas, and an elevated serum level of procollagen III peptide (PIIIP). We report the first case of a collagenofibrotic glomerulopathy patient who received a kidney transplantation. She received the kidney transplantation at the age of 18 years and the post-operative course was uneventful with good renal function on immunosuppression, which consisted of methylprednisolone, tacrolimus and basiliximab. Although urinary protein was negative, the serum level of PIIIP gradually elevated which suggests new collagen production in the graft. These findings indicate that the recipient had a systemic factor that stimulated collagen production. To investigate the prognosis of collagenofibrotic glomerulopathy, we carried out a questionnaire survey on 14 patients at 9 hospitals. None of these patients had received a kidney transplantation. However, 7 already had end-stage renal failure. Ten years after diagnosis, the renal survival rate was 49%. This rate is lower than cited in previous reports.

Background. Macrophages constitute 38% to 60% of infiltrating cells during acute renal allograft rejection. Their contribution to tissue damage during acute rejection was; examined by depleting macrophages in a rat model. Methods. Lewis rats underwent bilateral nephrectomy and then received a Dark Agouti renal allograft and liposomal-clodronate, control phosphate-buffered saline liposomes, or saline intravenously (n=7 per group) on days 1 and 3 postsurgery. Grafts were harvested on day 5. Results. Liposomal-clodronate treatment resulted in a 70% reduction in blood ED1(+) monocytes and 60% reduction in intragraft ED1(+) macrophages (both P<0.01). Half of all remaining interstitial ED1(+) cells were undergoing apoptosis (terminal deoxynucleotide transferase-mediated dUTP nick-end labeling(+)/ED1(+)), and thus functional depletion of more than 75% of macrophages was achieved. Histologic and functional parameters of acute rejection were attenuated: interstitial infiltrate, tubulitis, and glomerulitis (P<0.01); tubular cell apoptosis (P<0.001); tubular cell proliferation (P<0.01); and serum creatinine (P<0.01). Production of inducible nitric oxide synthase by infiltrating cells and urinary nitric oxide excretion was reduced by 90% (P<0.001). In contrast, no reduction in the number of other leukocytes was seen (CD3(+), CD4(+), CD8(+), and natural killer cells). Activation of lymphocytes (CD25(+)) and production of lymphocyte effector molecules (granzyme B) were unaltered. Conclusion. This study demonstrates that macrophages contribute to tissue damage during acute rejection.

Background. In vitro studies suggest that activation of the extracellular signal-regulated kinase (ERK) pathway plays a critical role in the proliferation of tubular epithelial and myofibroblast-like cells. However, little is known of ERK activation in individual cell types in normal or diseased kidney. The aims of this study were to (1) localize ERK activation within the kidney, and (2) examine the relationship between ERK activation and cell proliferation in the injured kidney. Methods. Unilateral ureteric obstruction (UUO) was induced in groups of six Wistar rats, which were killed at 30 minutes, 6 hours, and 1, 4, or 7 days after obstruction. Activation of ERK was identified using antibodies specific for the phosphorylated form of ERK (pERK) in Western blots and immunostaining. Proliferating cells were detected using bromodeoxyuridine (BrdU). Results. Western blotting showed abundant expression of the two ERK isoforms, ERK-1 and ERK-2, in normal rat kidney. Low levels of activated ERK (pERK-2 > pERK-1) were detected in normal rat kidney by Western blotting. Immunostaining showed that ERK activation in normal kidney was largely restricted to collecting ducts in the outer medulla. Within 30 minutes of ureter obstruction, Western blotting showed a sixfold increase in ERK activation followed by a second peak (14-fold increase) on days 4 and 7. The initial peak of ERK activation was localized to medullary collecting ducts and the thick ascending limb of Henle (TALH), whereas the second peak corresponded to a progressive increase in ERK activation in dilated collecting ducts and in interstitial cells in the cortex. Proliferation of tubular epithelial cells closely followed the pattern of ERK activation, being evident first in medullary collecting ducts and TALH on day 1, and then in cortical collecting ducts from day 4. Conclusion. This study has identified a discrete pattern of ERK activation in normal rat kidney and an increase in ERK activation following obstruction. The temporal and spatial relationship in which ERK activation preceded tubular cell proliferation suggest that ERK signaling plays a key role in tubular epithelial cell proliferation in the injured kidney.

Macrophages have been implicated in causing renal injury in both human and experimental kidney disease. The aim of the current study was to determine whether modulating the state of macrophage activation directly affects the capacity of these cells to cause renal injury. This was investigated using an adoptive transfer model in which macrophage activation can be manipulated in vitro, using interferon-gamma (IFN-gamma) or dexamethasone (Dex), and then macrophage-mediated renal injury determined in vivo. In this model, rats were made leukopenic by administration of cyclophosphamide (CyPh). Two days later (day 0), animals were injected with sheep anti-GBM serum followed by a single injection of rat NR8383 macrophages on day I and then killed 3 or 24 In after cell transfer. NR8383 macrophages were incubated IFN-gamma and/or Dex before adoptive transfer into animals. Induction of proteinuria and glomerular cell proliferation (PCNA+ cells) in this model was dependent on transfer of NR8383 macrophages. Exposure of macrophages to IFN-gamma for 18 h (but not 3 h) before transfer caused a twofold increase in the degree of proteinuria and glomerular cell proliferation compared with unstimulated cells (Nil versus IFN-gamma; P < 0.001). This was due to an increase in the number of transferred macrophages within the glomerulus and a significant increase in degree of renal injury per transferred glomerular macrophage. IFN-gamma increased iNOS and PDGF-B gene expression and upregulated adhesion molecule expression in NR8383 macrophages. In contrast, exposure of NR8383 cells to Dex for 18 h (but not 1 h) abrogated renal injury due to a failure of transferred macrophages to accumulate within the glomerulus. In addition, Dex abrogated renal injury caused by IFN-gamma-stimulated macrophages. In conclusion, activation of macrophages by IFN-gamma, independent of any effect on other leukocytes or renal cells, can substantially augment macrophage-mediated renal injury. This IFN-gamma augmentation of renal injury is sensitive to the action of glucocor-ticoids, which act directly on macrophages to prevent their recruitment to the inflamed glomerulus. This study provides the first evidence that it is possible to directly modulate macrophage-mediated renal injury.

Macrophages have been implicated in causing renal injury in both human and experimental kidney disease. The aim of the current study was to determine whether modulating the state of macrophage activation directly affects the capacity of these cells to cause renal injury. This was investigated using an adoptive transfer model in which macrophage activation can be manipulated in vitro, using interferon-gamma (IFN-gamma) or dexamethasone (Dex), and then macrophage-mediated renal injury determined in vivo. In this model, rats were made leukopenic by administration of cyclophosphamide (CyPh). Two days later (day 0), animals were injected with sheep anti-GBM serum followed by a single injection of rat NR8383 macrophages on day I and then killed 3 or 24 In after cell transfer. NR8383 macrophages were incubated IFN-gamma and/or Dex before adoptive transfer into animals. Induction of proteinuria and glomerular cell proliferation (PCNA+ cells) in this model was dependent on transfer of NR8383 macrophages. Exposure of macrophages to IFN-gamma for 18 h (but not 3 h) before transfer caused a twofold increase in the degree of proteinuria and glomerular cell proliferation compared with unstimulated cells (Nil versus IFN-gamma; P < 0.001). This was due to an increase in the number of transferred macrophages within the glomerulus and a significant increase in degree of renal injury per transferred glomerular macrophage. IFN-gamma increased iNOS and PDGF-B gene expression and upregulated adhesion molecule expression in NR8383 macrophages. In contrast, exposure of NR8383 cells to Dex for 18 h (but not 1 h) abrogated renal injury due to a failure of transferred macrophages to accumulate within the glomerulus. In addition, Dex abrogated renal injury caused by IFN-gamma-stimulated macrophages. In conclusion, activation of macrophages by IFN-gamma, independent of any effect on other leukocytes or renal cells, can substantially augment macrophage-mediated renal injury. This IFN-gamma augmentation of renal injury is sensitive to the action of glucocor-ticoids, which act directly on macrophages to prevent their recruitment to the inflamed glomerulus. This study provides the first evidence that it is possible to directly modulate macrophage-mediated renal injury.

Background. Glomerular macrophage accumulation is a feature of proliferative human and experimental glomerulonephritis. However, our understanding of the role of macrophages in the induction of renal injury is based upon indirect evidence from depletion studies, most of which lack specificity for this cell type. Therefore, an adoptive transfer approach was used to directly assess the potential of macrophages to induce renal injury. Methods. Accelerated anti-glomerular basement membrane (anti-GBM) disease was induced in rats by immunization with sheep IgG (day -5), followed by administration of sheep anti-rat GBM serum (day 0), with animals killed on day 2. To facilitate the adoptive transfer studies, immunized animals were made leukopenic by cyclophosphamide (CyPh) given on day -2. Bone marrow-derived (BM) or NR8383 macrophages were transferred by tail vein injection 24 hours after injection of anti-GBM serum, with animals killed 3 or 24 hours after transfer. Results. Pretreatment with CyPh prevented glomerular leukocyte accumulation and completely inhibited proteinuria, glomerular cell proliferation and hypercellularity in accelerated anti-GBM disease. Adoptive transfer led to significant glomerular accumulation of BM or NR8383 macrophages within 3 hours of injection, and this was still evident 24 hours later. Adoptive transfer of BM or NR8383 macrophages induced proteinuria (63 +/- 16 BM vs. 5 +/- 2 mg/24 h CyPh control; P < 0.001), glomerular cell proliferation (5.1 +/- 1.2 BM vs. 0.5 +/- 0.1 PCNA+ cells/gcs CyPh; P < 0.001) and glomerular hypercellularity (51.2 +/- 2.0 BM vs. 41.9 +/- 0.9 nuclei/gcs CyPh; P < 0.001). The degree of renal injury correlated with the number of transferred glomerular macrophages. Two-color immunostaining demonstrated that most glomerular proliferative cell nuclear antigen+ (PCNA+) proliferating cells were OX-7+ mesangial cells. CyPh treatment did not prevent up-regulation of glomerular intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule (VCAM-1) expression or an increase in urinary monocyte chemoattractant protein-1 (MCP-1) excretion. Conclusion. This study provides the first direct evidence that macrophages can induce renal injury in terms of proteinuria and mesangial cell proliferation.

Background. We established the reversible and the prolonged models of mesangial proliferative glomerulonephritis (GN) with anti-Thy 1 antibody 1-22-3. However, the essential factors leading to the prolonged glomerular alterations have not been identified. Methods. The expressions of several chemokines and cytokines were compared in the reversible and the prolonged models. Expression of fractalkine and the number of the fractalkine receptor CX3 CR1-positive cells in the glomeruli in the prolonged model were significantly higher than those in the reversible model. Then, the localization of fractalkine and the characteristics of CX3 CR1(+) cells were analyzed in glomeruli. To elucidate the significance of the fractalkine expression, we analyzed the expression in the model treated with angiotensin II receptor antagonist, candesartan. Results. Immunostaining of fractalkine was detected on endothelial cells on the fifth day, and fractalkine staining also was detected in the mesangial area on day 14. Major parts of the CX3 CR1(+) cells in the glomeruli were macrophages, especially ED3(+) cells. Candesartan treatment ameliorated the glomerular morphological findings at six weeks after disease induction. Although the treatment did not ameliorate the morphological finding at two weeks, decreased expression of fractalkine and CX3 CR1(+) were already detected at two weeks in rats treated with candesartan. Conclusions. Fractalkine expression and the recruitment of CX3 CR1(+) cells in glomeruli might play an important role in the development of the prolonged disease. These expressions could be predictors of the prolonged disease of the mesangial proliferative glomerulonephritis.

Background. We have previously reported that CD4 T lymphocytes and their cytokines contribute to development of Thy 1.1 glomerulonephritis (GN). FK506 is reported to suppress the production of Th1 cytokines. The aims of this study were to elucidate the role of Th1 cytokines on mesangial alteration and to examine whether FK506 is available for therapy of mesangial proliferative GN. Methods. The effects of daily treatments of FK506 from day -5 and from day +1 of Thy 1.1 GN induction on glomerular alterations were analyzed. Results. FK506 treatment with 1.0 and 0.3 mg/kg body weight (BW) daily from day 1 to day 4 significantly reduced the glomerular expression of mRNA for interferon-gamma (IFN-gamma; 1.0 mg/kg BW FK506, 32.4% to the placebo group, P < 0.01) and IL-2 (55.6%, P < 0.01) on day 5. FK506 treatment from day -5 of GN induction reduced proteinuria and glomerular alteration in a dose-dependent manner. Although no side effects were detected in rats with 0.3 mg/kg BW of FK506 treatment from day +1, the treatment also ameliorated proteinuria (day 14, 3.7 +/- 0.89 vs. 19.8 +/- 12.3 mg/100 g BW/day P < 0.05) and glomerular alterations [total cell number, 63.1 +/- 3.1 vs. 80.2 +/- 7.4, P < 0.01; matrix expansion, 0.90 +/- 0.30 vs. 1.34 +/- 0.27, P < 0.05; α-smooth muscle actin (αSMA) expression; 1.20 +/- 0.12 vs. 1.96 +/- 0.29, P < 0.01] on day 14. Conclusion. Th1 cytokines may play an important role in the development of mesangial proliferative glomerulonephritis, and could be targets for therapy. FK506 might be available for clinical use.

尿中の糸球体上皮細胞(podocyte)排泄の推移を観察し,管外性病変の進行を予測できるかを検討した.小児期発症IgA腎症20例(男児9例,女児11例・発症時年齢4〜15歳)について2年以上観察した結果,経過観察中に排泄が消失した群(A群,6例),持続して排泄が見られた群(B群,8例),排泄がほとんどなかった群(C群,6例)の3群が観察された.観察期間中に経皮的腎生検が行われた中で,病理組織学的に管外性病変の進行が確認された4例はいずれもB群であり,IgA腎症において尿中のpodocyte排泄が持続している症例では,糸球体管外性病変が進行している可能性があると考えられた

Background. From the observations of morphology seen in early phases of the experimental models of the irreversible mesangial proliferative glomerulonephritis, we hypothesized that podocyte injury is one of the important factors in bringing upon irreversible glomerular alterations. To verify this hypothesis, we investigated whether podocyte injury induced by puromycin aminonucleoside (PAN) injection affects the mesangial alterations of anti-Thy 1.1 glomerulonephritis. Methods. Female Wistar rats were injected with 0.5 mg monoclonal antibody (mAb) 1-22-3 five days after the injection of 10 mg or 5 mg/100 g body weight (BW) of puromycin aminonucleoside (PAN), and sacrificed at 7 days or 8 weeks after the mAb 1-22-3 injection. Results. Consecutive injections of 10 mg/100 g BW of PAN and mAb 1-22-3 caused the irreversible mesangial alteration with persistent proteinuria (at week 8, proteinuria 100.3 ± 57.8 mg/24 h, matrix score 1.13 ± 0.52, collagen type I score 2.04 ± 0.53, mRNA for collagen type I 227 ± 79% to the group with a single injection of 1-22-3). Although single injection of 5 mg/100 g BW of PAN was not capable of inducing abnormal proteinuria, consecutive injections of 5 mg/100 g BW of PAN and mAb 1-22-3 also caused irreversible mesangial alteration and persistent proteinuria. Conclusions. Podocyte injury might be an important factor that exacerbates mesangial proliferation and mesangial matrix expansion. The irreversible mesangial alterations caused by consecutive injections of PAN and mAb 1-22-3 may be a novel model that could be used to analyze the mechanism of progressive mesangial alteration.

The aim of this study was to evaluate whether the infiltrating T-lymphocyte can be a predictor in the disease progression of IgA nephropathy (IgAN). Twenty children with IgAN, followed for more than 5 years, were divided into progressive (n=5) and non-progressive groups (n=15). We assessed glomerular and interstitial infiltration of T-lymphocytes (CD4+ and CD8+ cells) and expression of α-smooth muscle actin (α-SMA) and transforming growth factor- β (TGF- β) using an indirect immunofluorescence method on the renal biopsies. We analyzed their relationship to the degree of proteinuria, histological changes, and prognosis. The number of CD8+ cells in glomeruli and in interstitium was higher in the progressive group than in the non-progressive group. The glomerular α-SMA staining was more intensive in the progressive group than in the non-progressive group. Urinary protein and the degree of histological changes were also higher in the progressive group than in the non-progressive group. Among these markers, the number of glomerular CD8+ cells was the most apparent difference between the two groups. In conclusion, these results indicate that the number of glomerular CD8+ cells is the most sensitive predictor of disease progression in childhood IgAN.

Background: Increased numbers of lymphocytes have been identified in biopsy specimens of human mesangial proliferative glomerulonephritis (GN). However, the causal relationship between infiltrating T lymphocytes and mesangial changes in mesangial proliferative GN has not been previously evaluated. In this study, we elucidated the role of lymphocytes in the development of mesangial proliferative GN. Method: Immunohistological and flow cytometric analyses as well as a reverse transcription-polymerase chain reaction (RTPCR) studies were performed in monoclonal antibody (mAb) 1-22-3- induced Thy 1.1 GN. To elucidate the role of these lymphocytes, depletion studies were carried out using anti-CD8 mAb (OX-8), which depletes both CD8+ T lymphocytes and natural killer (NK) cells and anti-CD5 mAb (OX-19), which depletes both CD4+ and CD8+ T lymphocytes. Results: Immunofluorescence (IF) studies revealed that NK cells and CD4+ T lymphocytes were recruited into glomeruli. Glomerular mRNA expression for interferon-γ interleukin-2 (IL-2), IL-10, and perforin increased after induction of GN. Increased expressions of several chemokines, which have the potential to attract lymphocytes, were also detected. Anti-CD8 mAb treatment completely prevented the recruitment of NK cells: however, it had no protective effect on proteinuria and mesangial injury. By contrast, anti-CD5 mAb treatment suppressed the recruitment of CD4+ T lymphocytes into glomeruli and reduced proteinuria (60.4 ± 25.7 vs. 120.0 ± 32.3 mg/day, P &lt 0.05) and mesangial changes evaluated by total number of cells in glomeruli (63.2 ± 6.0 vs. 81.4 ± 5.9, P &lt 0.01) and α- smooth muscle actin staining score (1.4 ± 0.2 vs. 2.2 ± 0.4, P &lt 0.01) on day 14 after induction of GN. mRNA expression for IL-2 was significantly reduced by OX-19 treatment. Conclusion: T lymphocytes participate in the development of mesangial proliferative GN.

低身長を主訴に来院し,染色体検査で14/X転座を認め,高精度分染法及びFISH法による解析で核型が46,X,der(X)t(X;14)(p22.13;q24.3)と診断された12歳女児を報告した.本例は精神発達遅滞,低身長及び出生時から多指症や口蓋裂等の表現型異常が認められ,14q遠位部トリソミーとしての症状が発現したものと考えられた.しかし,末梢血リンパ球及び皮膚線維芽細胞を用いたRBG法による分析の結果,観察された全ての細胞において派性X染色体は後期複製パターンを示し,不活化していると考えられ,又,転座した14q部分も後期複製を示したことから,不活化の影響は転座した14q部分にも及んでいると考えられた

心臓カテーテルによる大動脈造影所見を用いて正常小児の腎動脈径を計測し,正常値について検討した.大動脈径,年齢,身長,体重,体表面積,心拍出量及び心係数との相関について検討した結果,心係数を除く各因子との間に有意な相関が認められ,それぞれの回帰式が得られた.これらの回帰式を用いることによって非侵襲的に腎動脈の直径の推定が可能である