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Current study is a case report of a 5-year-old patient with T-cell acute lymphoblastic leukemia (ALL) and multiple urolithiasis. Complex factors, including glucocorticoid-induced hypercalciuria, fluid restriction for the syndrome of inappropriate secretion of antidiuretic hormone, and long-term bed rest, predispose children with ALL to develop urolithiasis. To prevent urinary urolithiasis formation, urinary calcium excretion should be monitored during chemotherapy and when administering glucocorticoids.

It is desirable that noninvasive differential diagnosis takes place without lymph node biopsy for histiocytic necrotizing lymphadenitis (HNL) or malignant lymphoma (ML). In this study, we propose a novel scoring model for the differential diagnosis of these diseases using clinical information and clinical findings. We retrospectively analyzed the data from 15 HNL and 13 ML pediatric patients. First, a univariate analysis identified 14 clinical factors with significant differences. Second, a subsequent analysis using receiver operating characteristic (ROC) curve analysis identified three factors among them with area under the ROC curve values of >0.95: body temperature (°C), maximum lymph node size (cm), and serum β2-microglobulin level (mg/L). Finally, the cut-off values of each of these three factors were determined and examined for the 28 cases. All 15 HNL cases were within 2–3 of the cut-off values among the three factors, no ML case was within two or more cut-off values. Thus, the diagnostic sensitivity and specificity of this novel scoring system were both 100%, indicating that clinical scoring with body temperature, maximum lymph node size, and β2-microglobulin are useful for distinguishing between HNL and ML.

The exact incidence of acute kidney injury (AKI) during chemotherapy for acute lymphoblastic leukemia (ALL)/lymphoblastic lymphoma (LBL) is unknown. Furthermore, childhood cancer survivors are at risk of AKI-chronic kidney disease transition. Thus, early diagnosis of AKI is crucial. This study aimed to elucidate the incidence of AKI in patients undergoing chemotherapy for pediatric ALL/LBL and to compare the usefulness of serum cystatin C (CysC)- and creatinine (Cr)-based estimated glomerular filtration rate (eGFR) as diagnostic measures. Data of 16 patients with ALL/LBL treated with a total of 75 courses of chemotherapy were retrospectively analyzed. CysC- and Cr-based eGFR were measured before and three times per week during therapy. To calculate the eGFR, an equation for Japanese children was used. AKI was diagnosed when eGFR dropped by ≥ 25% from the highest eGFR value obtained during the latest 2 weeks since the start of chemotherapy. AKI was graded based on the pediatric Risk, Injury, Failure, Loss, End Stage Renal Disease scale. All patients developed AKI during chemotherapy; however, more than 90% of the cases were mild and eventually recovered. No significant differences were found in the incidence of AKI between CysC- and Cr-based eGFR (p = 0.104). The median time to AKI diagnosis was significantly shorter in the CysC-based eGFR than in the Cr-based eGFR (8 vs. 17 days, p < 0.001). In this study, all patients with pediatric ALL/LBL could develop mild AKI during treatment. CysC-based eGFR is a more effective measure than Cr-based eGFR for the early diagnosis of AKI.

Background We describe a continuous monitoring method aimed at preserving nerve function during biopsy of lesions on the oculomotor nerve using stimulation of the oculomotor nerve proximal to the lesion. Case Description A 5-year-old girl with a recurrent left oculomotor nerve palsy and contrast-enhancing left oculomotor nerve mass on magnetic resonance imaging underwent a biopsy of the lesion to aid in its diagnosis. At the time of surgery, needle electrodes were inserted into the superior and inferior rectus muscles percutaneously, and cotton-covered electrodes were implanted into the oculomotor nerve proximal to the lesion. Compound muscle action potentials of the oculomotor nerve were measured continuously by monopolar stimulation. The lesion was mapped by direct stimulation, and the unresponsive area was excised. The amplitude of the compound muscle action potentials decreased during the resection but recovered postoperatively. After resection of the lesion, the compound muscle action potentials remained the same as they were preoperatively. No obvious postoperative oculomotor nerve palsy was observed. Conclusions This method of continuous monitoring of the function of the oculomotor nerve is simple to use and is suitable for lesions in close proximity to the oculomotor nerve.

The skin is responsible for a variety of physiological functions and is critical for wound healing and repair. Therefore, the regenerative capacity of the skin is important. However, stem cells responsible for maintaining the acral epithelium had not previously been identified. In this study, we identified the specific stem cells in the acral epithelium that participate in the long-term maintenance of sweat glands, ducts, and interadnexal epidermis and that facilitate the regeneration of these structures following injury. Lgr6-positive cells and Bmi1-positive cells were found to function as long-term multipotent stem cells that maintained the entire eccrine unit and the interadnexal epidermis. However, while Lgr6-positive cells were rapidly cycled and constantly supplied differentiated cells, Bmi1-positive cells were slow to cycle and occasionally entered the cell cycle under physiological conditions. Upon irradiation-induced injury, Bmi1-positive cells rapidly proliferated and regenerated injured epithelial tissue. Therefore, Bmi1-positive stem cells served as reservoir stem cells. Lgr5-positive cells were rapidly cycled and maintained only sweat glands; therefore, we concluded that these cells functioned as lineage-restricted progenitors. Taken together, our data demonstrated the identification of stem cells that maintained the entire acral epithelium and supported the different roles of three cellular classes.

The murine intestine, like that of other mammalians, continues to develop after birth until weaning; however, whether this occurs in response to an intrinsic developmental program or food intake remains unclear. Here, we report a novel system for the allotransplantation of small intestine and colon harvested from Lgr5 EGFP-IRES-CreERT2/+; Rosa26 rbw/+ mice immediately after birth into the subrenal capsule of wild-type mice. By histological and immunohistochemical analysis, the developmental process of transplanted small intestine and colon was shown to be comparable with that of the native tissues: mature intestines equipped with all cell types were formed, indicating that these organs do not require food intake for development. The intestinal stem cells in transplanted tissues were shown to self-renew and produce progeny, resulting in the descendants of the stem cells occupying the crypt-villus unit of the small intestine or the whole crypt of the colon. Collectively, these findings indicate that neonatal intestine development follows an intrinsic program even in the absence of food stimuli.

Although the existence of cancer stem cells in intestine tumors has been suggested, direct evidence has not been yet provided. Here, we showed, using the multicolor lineage-tracing method and mouse models of intestinal adenocarcinoma and adenoma that Bmi1- or Lgr5- positive tumorigenic cells clonally expanded in proliferating tumors. At tumor initiation and during tumor propagation in the colon, the descendants of Lgr5-positive cells clonally proliferated to form clusters. Clonal analysis using ubiquitous multicolor lineage tracing revealed that colon tumors derived from Lgr5-positive cells were monoclonal in origin but eventually merged with neighboring tumors, producing polyclonal tumors at the later stage. In contrast, the origin of small intestine tumors was likely polyclonal, and during cancer progression some clones were eliminated, resulting in the formation of monoclonal tumors, which could merge similar to colon tumors. These results suggest that in proliferating intestinal neoplasms, Bmi1- or Lgr5-positive cells represent a population of cancer stem cells, whereas Lgr5-positive cells also function as cells-of-origin for intestinal tumors.

Asingle cells in undifferentiated spermatogonia are considered to be the most primitive forms of germ stem cells (GSCs). Although GFRα1 is thought to be a marker of Asingle cells, we found that Bmi1(High) is more specific than GFRα1 for Asingle cells. Bmi1(High) expression in Asingle cells is correlated with seminiferous stages, and its expression was followed by the proliferative stage of Asingle GSCs. In contrast, GFRα1 expression was seminiferous stage-independent. Fate analyses of EdU-positive Bmi1(High)-positive cell-derived Asingle cells revealed that these cells self-renewed or generated transient amplifying Apaired cells. Bmi1(High)-positive cells were resistant to irradiation-induced injury, after which they regenerated. Elimination of Bmi1(High)-positive cells from seminiferous tubules resulted in the appearance of tubules with seminiferous stage mismatches. Thus, in this study, we found that Bmi1(High) is a seminiferous stage-dependent marker for long-term GSCs and that Bmi1(High)-positive cells play important roles in maintaining GSCs and in regenerating spermatogenic progenitors after injury.

Despite the strong need for the establishment of a lingual epithelial cell culture system, a simple and convenient culture method has not yet been established. Here, we report the establishment of a novel lingual epithelium organoid culture system using a three-dimensional matrix and growth factors. Histological analyses showed that the generated organoids had both a stratified squamous epithelial cell layer and a stratum corneum. Very recently, we showed via a multicolor lineage tracing method that Bmi1-positive stem cells exist at the base of the epithelial basal layer in the interpapillary pit. Using our new culture system, we found that organoids could be generated by single Bmi1-positive stem cells and that in the established organoids, multiple Bmi1-positive stem cells were generated at the outermost layer. Moreover, we observed that organoids harvested at an early point in culture could be engrafted and maturate in the tongue of recipient mice and that the organoids generated from carcinogen-treated mice had an abnormal morphology. Thus, this culture system presents valuable settings for studying not only the regulatory mechanisms of lingual epithelium but also lingual regeneration and carcinogenesis.

Abstract 4585 Malignant infantile osteopetrosis is an autosomal recessive disease characterized by a lack of osteoclastic function with incidence of 1: 200,000 to 1: 300,000. In consequence of disturbed bone building and remodeling, affected patients have osteosclerosis, dense fragile bone, and a marked reduction in the bone marrow cavity. Clinical features, such as anemia, thrombocytopenia, hepatosplenomegaly, bone fractures, bone deformity, and cranial nerve entrapment, appear soon after birth or within the first years of life. In the natural course of the disease, only 30% of children can survive to more than six years old. Because osteoclasts are of hematopoietic origin, allogeneic stem cell transplantation is the only curable therapy. Recently, successful HLA-haploidentical hematopoietic stem cell transplantation (HSCT) has been reported for patients without HLA- matched donors. We also present an infant with malignant infantile osteopetrosis who underwent HLA-haploidentical HSCT. A four-month-old boy was referred to our hospital for splenomegaly and pancytopenia. A physical examination revealed failure to thrive, distention of anterior fontanel, and hepatosplenomegaly. Laboratory findings indicated thrombocytopenia and anemia, and elevated alkaline phosphatase. Osteoclasts were hyperplastic in bone marrow. Radiography showed homogeneous and sclerotic bones with absence of corticomedullary junction, and bone marrow scintigraphy showed no accumulation of bone marrow. Subsequently, TCIRG1 mutations are identified. All these findings are compatible with malignant infantile osteopetrosis and the diagnosis was made. We conducted HSCT to revive his osteoclastic function. However, he didn't have HLA-matched donors not only in relatives but also in the bone marrow bank and the cord blood bank of Japan. We attempted HLA-haploidentical bone marrow transplantation (BMT) from his father with written informed consent. The patient was prepared for BMT at 11 months of age by conditioning with busulfan (6 mg/kg × 4/day × 4 days), cyclophosphamide (360mg/m2 × 2 days) and antithymocyte globulin (2 mg/kg/day × 3 days). Prophylaxis for graft-versus-host disease (GVHD) included tacrolimus and short-term methotrexate. Bone marrow nuclear cells (1×109/kg) were collected from his father and transplanted without any manipulations. The engraftment of neutrophils was confirmed on day 9 after BMT. Acute GVHD was limited to the skin (grade I). At 2 months after BMT, neutrophil counts, platelet counts, and hemoglobin level were all within normal limits. Bone marrow scintigraphy revealed a significant uptake in bone marrow. There have been no chronic GVHD up to six months after BMT.HSCT for patients with osteopetrosis is challenging as it is reported that HLA-haploidentical HSCT has the high rate of acute and chronic GVHD. Fortunately, HLA-haploidentical BMT was successful in our patient. This might be due to our regimen for prophylaxis of rejection and GVHD. We used ATG in conjunction with tacrolimus: this regimen may be prophylactic not only for rejection but also for GVHD as previously reported (Schulz et al. Blood 2002; 99: 3458–3460). In conclusion, HLA-haploidentical family donors should be considered as the alternative hematopoietic stem cell source for patients with malignant infantile osteopetrosis in condition without HLA-matched donors. Disclosures: No relevant conflicts of interest to declare.