Green-Sensitive, Long-Lived, Step-Functional Anion Channelrhodopsin-2 Variant as a High-Potential Neural Silencing Tool.

Anion channelrhodopsin-2 (GtACR2) was identified from the alga Guillardia theta as a light-gated anion channel, providing a powerful neural silencing tool for optogenetics. To expand its molecular properties, we produced here GtACR2 variants by strategic mutations on the four residues around the retinal chromophore (i.e., R129, G152, P204, and C233). After the screening with the Escherichia coli expression system, we estimated spectral sensitivities and the anion channeling function by using the HEK293 expression system. Among the mutants, triple (R129M/G152S/C233A) and quadruple (R129M/G152S/P204T/C233A) mutants showed the significantly red-shifted absorption maxima (λmax = 498 and 514 nm, respectively) and the long-lived channel-conducting states (the half-life times were 3.4 and 5.4 s, respectively). In addition, both mutants can be activated and inactivated by different wavelengths, representing their step-functional ability. We nicknamed the quadruple mutant "GLaS-ACR2" from its green-sensitive, long-lived, step-functional properties. The unique characteristics of GLaS-ACR2 suggest its high potential as a neural silencing tool.

Polyarteritis Nodosa with Marked Eosinophilia, Associated with Severe Gastrointestinal Tract Involvement and Recurrent Venous Thrombosis.

A 45-year-old man was admitted with acute abdominal pain and eosinophilia. Abdominal computed tomography revealed thickness of the ascending and transverse colon with decreased contrast enhancement and a small amount of ascites. In an emergency operation, the necrotic colon was resected. Histopathology showed subserous medium-sized arteritis with abundant eosinophil infiltrates and thrombosis in the portal vein branches. He was diagnosed with polyarteritis nodosa (PAN), and immunosuppressive therapy improved his condition. Two years later, the disease recurred with ischemic cutaneous lesions and marked eosinophilia. Our experience suggests that marked eosinophilia in PAN may imply severe organ involvement, including gastrointestinal necrosis, as well as the association of venous thrombosis.

Mutational analysis of the conserved carboxylates of anion channelrhodopsin-2 (ACR2) expressed in Escherichia coli and their roles in anion transport.

Anion channelrhodopsin-2 (ACR2), a light-gated channel recently identified from the cryptophyte alga Guillardia theta, exhibits anion channel activity with exclusive selectivity. In addition to its novel function, ACR2 has become a focus of interest as a powerful tool for optogenetics. Here we combined experimental and computational approaches to investigate the roles of conserved carboxylates on the anion transport activity of ACR2 in Escherichia coli membrane. First, we replaced six conserved carboxylates with a neutral residue (i.e. E9Q, E56Q, E64Q, E159Q, E219Q and D230N), and measured anion transport activity using E. coli expression system. E159Q and D230N exhibited significantly lower anion transport activity compared with wild-type ACR2 (1/12~1/3.4), which suggests that E159 and D230 play important roles in the anion transport. Second, to explain its molecular aspects, we constructed a homology model of ACR2 based on the crystal structure of a cation channelrhodopsin (ChR). The model structure showed a cavity formed by four transmembrane helices (TM1, TM2, TM3 and TM7) similar to ChRs, as a putative anion conducting pathway. Although E159 is not located in the putative pathway, the model structure showed hydrogen bonds between E159 and R129 with a water molecule. D230 is located in the pathway near the protonated Schiff base (PSB) of the chromophore retinal, which suggests that there is an interaction between D230 and the PSB. Thus, we demonstrated the functional importance and the hypothetical roles of two conserved carboxylates, E159 and D230, in the anion transport activity of ACR2 in E. coli membrane.

Unusual underlying disorder for pulmonary embolism: Cold agglutinin disease.

Cold agglutinin disease (CAD) is a form of autoimmune hemolytic anemia caused by cold-reacting autoantibodies. The manifestations of CAD are commonly anemia, acrocyanosis, and fatigue caused by hemolysis and agglutination of red blood cells (RBCs) at a temperature lower than normal body temperature. We report a case of CAD presenting with pulmonary embolisms in an 86-year-old man. The patient visited our emergency department complaining of acute chest pain and respiratory distress. Laboratory data showed decreased RBC and hematocrit and markedly elevated mean corpuscular hemoglobin (MCH) and MCH concentration (MCHC). A contrast-enhanced computed tomographic scan demonstrated bilateral massive pulmonary embolisms. After admission, diagnosis of CAD was made on the basis of a high cold agglutinin titer without other factors of coagulation. CAD can contribute to the onset of pulmonary embolisms. It is necessary to incubate blood samples at 37 °C when laboratory data show markedly elevated MCH and MCHC and to consider the presence of cold agglutinins as an underlying disorder for the formation of venous thrombosis. .

[Two cases of discordant lymphomas consisting of MALT lymphoma and follicular lymphoma].

We report here rare cases of discordant lymphoma consisting of MALT lymphoma and follicular lymphoma. Case 1: A 53-year-old woman was diagnosed with MALT lymphoma of the left parotid gland and follicular lymphoma of the duodenum and small intestine. Case 2: A 38-year-old woman was diagnosed with MALT lymphoma of the intestine and follicular lymphoma of the duodenum and bone marrow. Recently, it has been suggested that duodenal follicular lymphoma has intermediate characteristics of nodal follicular lymphoma and MALT lymphoma. It is interesting that both of these cases demonstrated duodenal follicular lymphoma. These cases suggest that MALT lymphoma and duodenal follicular lymphoma share some common pathological condition.

[Complication of topoisomerase II inhibitor-related acute promyelocytic leukemia with t(1;10) (q21;q26) in a patient with Sézary syndrome].

A 74-year-old man was diagnosed as having Sézary syndrome in 1999. Treatment with combination chemotherapy could not completely control both the erythroderma and Sézary cells. However, treatment with oral administration of etoposide was able to maintain the patient in a good condition for about 4 years. In June 2004, he developed topoisomerase II inhibitor-related acute promyelocytic leukemia. Chromosomal analysis demonstrated abnormalities of t(1;10) (q21;q26) and t(15;17) (q22;q12) in 17 of 20 cells. Despite treatment with ATRA and combination chemotherapy, the patient died of brain hemorrhage.

Use of imatinib mesylate for favorable control of hypercalcemia mediated by parathyroid hormone-related protein in a patient with chronic myelogenous leukemia at blast phase.

The case of a 72-year-old woman with chronic myelogenous leukemia in blast phase (BP) with hypercalcemia is reported. Bone x-ray examination revealed multiple osteolytic lesions throughout the body. The serum level of parathyroid hormone-related protein (PTHrP) was elevated, and PTHrP messenger RNA (mRNA) was detectable in the peripheral blood mononuclear cells (PBMNC) at BP but was not detectable at chronic phase (CP).Treatment with conventional chemotherapy did not completely control either serum calcium level or serum PTHrP level. Treatment with imatinib mesylate (imatinib) alone rapidly normalized these parameters in parallel with a decrease in the number of blast cells. The treatment also maintained the patient in good condition for approximately 3 months, even though the number of blast cells, serum calcium level, serum PTHrP level, and PTHrP mRNA level increased at the terminal stage. Mutations of the p53, K-Ras, and BCR-ABL genes in PBMNC at BP were absent. A noteworthy feature in this patient was that PBMNC at BP but not at CP showed high Lyn mRNA expression. Taken together the findings showed that production of PTHrP by blast cells was favorably controlled by imatinib therapy alone. Imatinib may prolong survival time at BP even though the patients have the complication of PTHrP-mediated hypercalcemia.