Expansion of Anticomplement Therapy Indications from Rare Genetic Disorders to Common Kidney Diseases.

Complement constitutes a major part of the innate immune system. The study of complement in human health has historically focused on infection risks associated with complement protein deficiencies; however, recent interest in the field has focused on overactivation of complement as a cause of immune injury and the development of anticomplement therapies to treat human diseases. The kidneys are particularly sensitive to complement injury, and anticomplement therapies for several kidney diseases have been investigated. Overactivation of complement can result from loss-of-function mutations in complement regulators; gain-of-function mutations in key complement proteins such as C3 and factor B; or autoantibody production, infection, or tissue stresses, such as ischemia and reperfusion, that perturb the balance of complement activation and regulation. Here, we provide a high-level review of the status of anticomplement therapies, with an emphasis on the transition from rare diseases to more common kidney diseases.

MASP3 Deficiency in Mice Reduces but Does Not Abrogate Alternative Pathway Complement Activity Due to Intrinsic Profactor D Activity.

Complement factor D (FD) is a rate-limiting enzyme of the alternative pathway (AP). Recent studies have suggested that it is synthesized as an inactive precursor and that its conversion to enzymatically active FD is catalyzed by mannan-binding lectin-associated serine protease 3 (MASP3). However, whether MASP3 is essential for AP complement activity remains uncertain. It has been shown that Masp1/3 gene knockout did not prevent AP complement overactivation in a factor H-knockout mouse, and a human patient lacking MASP3 still retained AP complement activity. In this study, we have assessed AP complement activity in a Masp3-knockout mouse generated by CRISPR/Cas9 editing of the Masp1/3 gene. We confirmed specific Masp3 gene inactivation by showing intact MASP1 protein expression and absence of mature FD in the mutant mice. Using several assays, including LPS- and zymosan-induced C3b deposition and rabbit RBC lysis tests, we detected plasma concentration-dependent AP complement activity in Masp3 gene-inactivated mice. Thus, although not measurable in 5% plasma, significant AP complement activity was detected in 20-50% plasma of Masp3 gene-inactivated mice. Furthermore, whereas FD gene deletion provided more than 90% protection of CD55/Crry-deficient RBCs from AP complement-mediated extravascular hemolysis, Masp3 gene deletion only provided 30% protection in the same study. We also found pro-FD to possess intrinsic catalytic activity, albeit at a much lower level than mature FD. Our data suggest that MASP3 deficiency reduces but does not abrogate AP complement activity and that this is explained by intrinsic pro-FD activity, which can be physiologically relevant in vivo.

The germ cell-specific TAP-like protein NXF-2 forms a novel granular structure and is required for tra-2 3'UTR-dependent mRNA export in Caenorhabditis elegans.

TAP is a general mRNA export receptor and is highly conserved among eukaryotes. The nematode Caenorhabditis elegans has another TAP-like protein, NXF-2, but little is known about its function. In this study, we show that NXF-2 is specifically expressed in germ cells and forms a novel granular structure that is different from that of P granules and that NXF-2 granules are anchored to the nuclear periphery in the mitotic region of the hermaphrodite gonad. In contrast, NXF-2 granules are released within the whole cytoplasm in the meiotic region, where the feminization gene tra-2 starts to function. Both inhibition of XPO-1 (an ortholog of the export receptor CRM1) and mutation of the nuclear export signal of NXF-2 caused the release of NXF-2 granules from the nuclear periphery, indicating that anchoring of NXF-2 granules depends on XPO-1 function. Moreover, inhibition of NXF-2 resulted in a substantial nuclear accumulation of the reporter mRNA carrying the tra-2 3'UTR. These results suggest that, together with XPO-1, NXF-2 exports and anchors tra-2 mRNA to the nuclear periphery to avoid precocious translation until the germ cells reach the meiotic region, thereby contributing to the regulation of tra-2 mRNA expression.

Risk score for predicting death from other causes after curative gastrectomy for gastric cancer.

BACKGROUND: The number of patients who die from causes other than gastric cancer after R0 resection is increasing in Japan, due in part to the aging population. However, few studies have comprehensively investigated the clinicopathological risks associated with deaths from other causes after gastrectomy. This study aimed to build a risk score for predicting such deaths. METHODS: We retrospectively reviewed clinical data for 3575 patients who underwent gastrectomy for gastric cancer at nine institutions in Japan between January 2010 and December 2014. RESULTS: The final study population of 1758 patients were assigned to Group A (n = 187): patients who died from other causes within 5 years of surgery, and Group B (n = 1571): patients who survived ≥ 5 years after surgery. Multivariate analysis identified nine characteristics as risk factors for poor survival: age ≥ 75 years, male sex, body mass index < 22 kg/m2, Eastern Cooperative Oncology Group Performance Status (≥ 1), diabetes mellitus, cardiovascular/cerebrovascular disease, other malignant diseases, preoperative albumin level < 3.5 g/dL, and total gastrectomy. Patients with risk scores of 0-2, 3-4, or 5-9 (based on 1 point per characteristics) were classified into Low-risk, Intermediate-risk, and High-risk groups, respectively. The 5-year survival rates were 96.5%, 85.3%, and 56.5%, for the Low-, Intermediate-, and High-risk groups, respectively, and the hazard ratio (95% confidence intervals) was 16.33 (10.85-24.58, p < 0.001) for the High-risk group. CONCLUSIONS: The risk score defined here may be useful for predicting deaths from other causes after curative gastrectomy.

A novel endovascular method of atherectomy for calcified common femoral and popliteal disease using the crosser system: Crossbow and Rambow techniques.

OBJECTIVES: This study aims to report the efficacy and safety of new atherectomy methods using the Crosser system for calcified lesions in the common femoral and popliteal artery: the Crosser system supported by bended 0.014 wire (Crossbow) technique and retrograde approach of sheathless Crosser system supported by bended 0.014 wire (Rambow) technique. MATERIALS AND METHODS: This report describes a single-center, retrospective study. A total of 23 patients (mean ± SD age, 73 ± 10 years; 19 men) with symptomatic peripheral artery disease received the Crossbow technique and Rambow technique for treatment of calcified common femoral and popliteal disease; these patients were enrolled between October 2013 and October 2015. The primary efficacy outcome was acute technical success, defined as achievement of residual stenosis < 30% for stenting and < 50% for angioplasty or atherectomy. The primary safety outcome was assessed on the basis of angiographic complications. RESULTS: The Crossbow and Rambow techniques were undertaken in 100% and 17% of the patients, respectively. Acute technical success was achieved in 96% of the patients. There were two embolic events. CONCLUSION: Crossbow and Rambow techniques could be effective atherectomy methods of calcified common femoral and popliteal disease. Regarding safety, embolic protection devices may be needed for our atherectomy methods.

Parallel Wiring Using a Single Intravascular Ultrasound Catheter with Double Rapid Exchange Lumens Technique for Chronic Total Occlusions in Peripheral Artery Disease.

PURPOSE: To describe a parallel wiring using a single intravascular ultrasound catheter with double rapid exchange lumens (PASSABLE) technique for peripheral CTOs. TECHNIQUE: The technique is demonstrated in a 73-year-old patient with CTOs of the superficial femoral and popliteal artery. Intravascular ultrasound (IVUS) examination revealed the first guidewire was advanced to the intramedial space of the popliteal artery. Following insertion of the first guidewire into only the distal rapid exchange lumen of the IVUS catheter and a second guidewire into the proximal rapid exchange lumen, a guidewire torquer was passed over it and tightened close to an exit port of the proximal rapid exchange lumen to prevent it from exiting an entry port while advancing the IVUS catheter. The IVUS catheter was advanced to the intraplaque region using only the distal rapid exchange lumen and the second guidewire was then advanced to the intraplaque region under IVUS guidance. The IVUS-guided wiring using the second guidewire on both the distal and proximal rapid exchange lumen was continued and resulted in a successful guidewire crossing into the distal true lumen. CONCLUSION: This novel technique may prove beneficial in enabling operators to perform IVUS-guided parallel wiring more easily and efficiently.

Prediction of Technical Failure of Inframalleolar Angioplasty in Patients with Chronic Limb Threatening Ischaemia.

OBJECTIVE: This study sought to: determine anatomically evaluated predictors of the technical failure of inframalleolar angioplasty (IMA), develop a predictive model for unsuccessful IMA, and investigate the effect of IMA on clinical outcomes in patients with chronic limb threatening ischaemia (CLTI). METHODS: This single centre retrospective observational study enrolled 159 patients with CLTI who underwent IMA for de novo occluded lesions between November 2017 and May 2021. These patients were divided into two groups: the Failed IMA group (n = 62) and the Successful IMA group (n = 97). RESULTS: In multivariable analysis, no target vessel outflow (OR 39.8, 95% CI 10.7 - 148, p < .001), medial artery calcification (MAC) grade (OR 4.91, 95% CI 1.40 - 17.3, p = .010), and occluded pedal arch (OR 5.2, 95% CI 1.2 - 22.7, p = .030) were identified as independent predictors of IMA technical failure. The risk prediction model had an area under the receiver operating characteristic curve (AUC) of 0.93; after bootstrapping adjustment for optimism, this value represented a corrected AUC of 0.95. The patients in the Successful IMA group had a significantly higher proportion of wound healing at 12 months than those in the Failed IMA group (log rank p = .030). IMA technical failure was associated with a significant change in the proportion of wound healing (HR 0.59, 95% CI 0.37- 0.94, p = .030). CONCLUSION: No target vessel outflow, MAC grade, and occluded pedal arch were independent predictors of IMA technical failure. Additionally, successful IMA was associated with better wound healing outcomes at 12 months. Furthermore, a model incorporating these three predictors precisely predicted IMA technical failure.

Hepatic metastasis of gastric cancer is associated with enhanced expression of ethanolamine kinase 2 via the p53-Bcl-2 intrinsic apoptosis pathway.

BACKGROUND: Gastric cancer (GC) with hepatic metastasis has a poor prognosis. Understanding the molecular mechanisms involved in hepatic metastasis may contribute to the development of sensitive diagnostic biomarkers and novel therapeutic strategies. METHODS: We performed transcriptome analysis of surgically resected specimens from patients with advanced GC. One of the genes identified as specifically associated with hepatic metastasis was selected for detailed analysis. GC cell lines with knockout of the candidate gene were evaluated in vitro and in vivo. Expression of the candidate gene was analysed in GC tissues from 300 patients. RESULTS: Ethanolamine kinase 2 (ETNK2) was differentially upregulated in GC patients with hepatic metastasis. ETNK2 expression was elevated in GC cell lines derived from haematogenous metastases. ETNK2 knockout significantly suppressed proliferation, invasion, and migration; increased apoptosis; reduced Bcl-2 protein expression; and increased phosphorylated p53 expression. In mouse xenograft models, ETNK2 knockout virtually abolished hepatic metastasis. Stratification of GC patients based on ETNK2 mRNA level revealed significant associations between high ETNK2 tumour expression and both hepatic recurrence and worse prognosis. CONCLUSIONS: Upregulation of ETNK2 in GC enhances hepatic metastasis, possibly via dysregulation of p53-Bcl-2-associated apoptosis. ETNK2 expression may serve as a biomarker for predicting hepatic recurrence and a therapeutic target.

G-protein subunit gamma-4 expression has potential for detection, prediction and therapeutic targeting in liver metastasis of gastric cancer.

BACKGROUND: The liver is the most common site for haematogenous metastasis of gastric cancer, and liver metastasis is fatal. METHODS: We conducted a transcriptomic analysis between metastatic foci in the liver, primary tumour and adjacent tissues from gastric cancer patients with metastasis limited to the liver. We determined mRNA expression levels in tumour tissues of 300 patients with gastric cancer via quantitative RT-PCR. The oncogenic phenotypes of GNG4 were determined with knockdown, knockout and forced expression experiments. We established and compared subcutaneous and liver metastatic mouse xenograft models of gastric cancer to reveal the roles of GNG4 in tumorigenesis in the liver. RESULTS: GNG4 was upregulated substantially in primary gastric cancer tissues as well as liver metastatic lesions. High levels of GNG4 in primary cancer tissues were associated with short overall survival and the likelihood of liver recurrence. Functional assays revealed that GNG4 promoted cancer cell proliferation, the cell cycle and adhesiveness. Tumour formation by GNG4-knockout cells was moderately reduced in the subcutaneous mouse model and strikingly attenuated in the liver metastasis mouse model. CONCLUSIONS: GNG4 expression may provide better disease monitoring for liver metastasis, and GNG4 may be a novel candidate therapeutic target for liver metastasis.

Non-Destructive and Quantitative Evaluation of Rebar Corrosion by a Vibro-Doppler Radar Method.

It is well known that evaluation of rebar corrosion is important for the maintenance of reinforced concrete structures, but, it is difficult to simply, quickly and quantitatively evaluate the amount of corrosion of rebars embedded in concrete by conventional non-destructive evaluation (NDE) methods such as electrical, electromagnetic and mechanical method. This paper proposes a vibro-Doppler radar (VDR) measurement method to quantitatively evaluate rebar corrosion by measuring the vibration ability of the rebar forcibly vibrated in concrete by an excitation coil. It is experimentally demonstrated in RC test pieces that the rebar vibration displacement obtained by developed VDR method is valid and is less affected by the moisture in the concrete. In addition, simultaneous monitoring of the rebar vibration displacement of the test pieces is performed through an electrolytic corrosion test and the measured vibration displacement is compared to the rebar corrosion loss evaluated. As the results, it is cleared that the rebar vibration displacement starts to increase from slightly before the occurrences of corrosion crack on the concrete surface as the corrosion loss increases. It is also shown that the rebar vibration displacement becomes 4 times higher than that in initial condition at the rebar corrosion loss of 250 mg/cm2. This implies that the VDR has potential to nondestructively and quantitatively evaluate rebar corrosion in concrete.

A Local Oscillator Phase Compensation Technique for Ultra-Wideband Stepped-Frequency Continuous Wave Radar Based on a Low-Cost Software-Defined Radio.

The paper discusses a way to configure a stepped-frequency continuous wave (SFCW) radar using a low-cost software-defined radio (SDR). The most of high-end SDRs offer multiple transmitter (TX) and receiver (RX) channels, one of which can be used as the reference channel for compensating the initial phases of TX and RX local oscillator (LO) signals. It is same as how commercial vector network analyzers (VNAs) compensate for the LO initial phase. These SDRs can thus acquire phase-coherent in-phase and quadrature (I/Q) data without additional components and an SFCW radar can be easily configured. On the other hand, low-cost SDRs typically have only one transmitter and receiver. Therefore, the LO initial phase has to be compensated and the phases of the received I/Q signals have to be retrieved, preferably without employing an additional receiver and components to retain the system low-cost and simple. The present paper illustrates that the difference between the phases of TX and RX LO signals varies when the LO frequency is changed because of the timing of the commencement of the mixing. The paper then proposes a technique to compensate for the LO initial phases using the internal RF loopback of the transceiver chip and to reconstruct a pulse, which requires two streaming: one for the device under test (DUT) channel and the other for the internal RF loopback channel. The effect of the LO initial phase and the proposed method for the compensation are demonstrated by experiments at a single frequency and sweeping frequency, respectively. The results show that the proposed method can compensate for the LO initial phases and ultra-wideband (UWB) pulses can be reconstructed correctly from the data sampled by a low-cost SDR.

[A Case of Effective Chemoradiotherapy for the Treatment of Distant Recurrence of Squamous Cell Carcinoma in the Anal Canal].

A 75-year-old woman diagnosed with squamous cell carcinoma of the anal canal was treated using chemoradiotherapy and revealed a complete response to the tumor. After 6 months of treatment, swollen para-aortic lymph nodes were found to develop. The patient received the same regimen of chemoradiotherapy again, resulting in lymph node disappearance. However, 2 months later, PET-CT revealed accumulation of FDG in the axillary and cervical lymph nodes. Chemoradiotherapy was performed for the third time. Swollen lymph nodes were found to disappear. After 12 months, para-aortic, axillary, and cervical lymph nodes developed, following which she received BSC; subsequently, she died after 38 months of the carcinoma diagnosis.

Therapeutic monoclonal antibody targeting of neuronal pentraxin receptor to control metastasis in gastric cancer.

BACKGROUND: Controlling metastasis is essential for improving the prognosis of patients with gastric cancer (GC). Here, we aimed to identify a molecule required for GC metastasis and to investigate its potential utility as a target for the development of therapeutic antibodies (Abs). METHODS: Transcriptome and bioinformatics analyses of human GC cell lines identified the neuronal pentraxin receptor (NPTXR) as a candidate molecule. NPTXR function was probed by modulating its expression in GC cells and assessing the effects on intracellular signaling and malignant behaviors in vitro and in mouse xenograft models. We also generated anti-NPTXR Abs and Nptxr-/- mice, and assessed the clinical significance of NPTXR expression in GC specimens. RESULTS: NPTXR mRNA expression in clinical specimens was associated with disease progression and was significantly higher in tissues from GC patients with distant metastasis compared with those without. NPTXR regulated expression of genes involved in metastatic behaviors as well as activation of the PI3K-AKT-mTOR, FAK-JNK, and YAP signaling pathways. NPTXR silencing promoted caspase-mediated apoptosis and attenuated GC cell proliferation, cell cycle progression, migration, invasion, adhesion, stem cell-like properties, and resistance to 5-fluorouracil in vitro, and also inhibited the tumorigenicity of GC cells in vivo. Anti-NPTXR Abs inhibited GC peritoneal metastasis in mice. Nptxr-/- mice showed no abnormalities in reproduction, development, metabolism, or motor function. CONCLUSIONS: NPTXR plays an essential role in controlling the malignant behavior of GC cells in vitro and in vivo. NPTXR-targeting Abs may thus have utility as novel diagnostic tools and/or treatment modalities for GC.

Fraser extracellular matrix complex subunit 1 promotes liver metastasis of gastric cancer.

Liver metastasis is often fatal in patients with gastric cancer, therefore, we aimed to identify genes associated with the mechanisms of liver metastasis of gastric cancer (GC) and to investigate their potential to predict recurrence and to serve as targets of therapy. Recurrence pattern-specific transcriptome analysis was performed to identify liver metastasis-associated genes. A stable knockout cell line was generated to investigate metabolic pathways that contribute to the malignant phenotype in vitro and vivo. Three hundred GC patients were analyzed to demonstrate an association between gene expression levels and clinicopathological parameters. As a results extracellular matrix complex subunit 1 (FRAS1) was identified as a liver metastasis-associated gene. Pathway analysis revealed that FRAS1 expression was significantly correlated with the expression of genes encoding TGFB1, MAP1B, AHNAK, BMP2, MUC1, BIRC5, MET, CDH1, RB1 and MKI67. FRAS1 expression was associated with the activation of the EGFR and PI3K signaling pathways. The proliferation ability of FRAS1 knockout cell line (FRAS1-KO) was inhibited compared to that of the parent cell line through caspase activity increment and cell cycle alteration. FRAS1-KO cells exhibited increased responsiveness to oxygen stress and diminished stemness, invasiveness, and migration. Mouse models of GC revealed decreases in tumor formation and generation of metastasis by FRAS1-KO cells. Moreover, the cumulative liver recurrence rate was significantly increased in patients with GC with high FRAS1 expression levels. We concluded that FRAS1 contributes to the malignant phenotype of GC, especially liver metastasis, and may therefore serve as a predictive marker or a target for treating liver metastasis.

MRG-1 is required for both chromatin-based transcriptional silencing and genomic integrity of primordial germ cells in Caenorhabditis elegans.

In Caenorhabditis elegans, germline cells remain transcriptionally silenced during embryogenesis. The transcriptional silencing is achieved by two different mechanisms: One is the inhibition of RNA polymerase II in P2-P4 cells at the establishment stage, and another is chromatin-based silencing in two primordial germ cells (PGCs) at the maintenance stage; however, the molecular mechanism underlying chromatin-based silencing is less understood. We investigated the role of the chromodomain protein MRG-1, which is an essential maternal factor for germline development, in transcriptional silencing in PGCs. PGCs lacking maternal MRG-1 showed increased levels of two histone modifications (H3K4me2 and H4K16ac), which are epigenetic markers for active transcription, and precocious activation of germline promoters. Loss of MES-4, a H3K36 methyltransferase, also caused similar derepression of the germline genes in PGCs, suggesting that both MRG-1 and MES-4 function in chromatin-based silencing in PGCs. In addition, the mrg-1 null mutant showed abnormal chromosome structures and a decrease in homologous recombinase RAD-51 foci in PGCs, but the mes-4 null mutant did not show such phenotypes. Taken together, we propose that MRG-1 has two distinct functions: chromatin-based transcriptional silencing and preserving genomic integrity at the maintenance stage of PGCs.

Complement Factor H Mutation W1206R Causes Retinal Thrombosis and Ischemic Retinopathy in Mice.

Single-nucleotide polymorphisms and rare mutations in factor H (FH; official name, CFH) are associated with age-related macular degeneration and atypical hemolytic uremic syndrome, a form of thrombotic microangiopathy. Mice with the FH W1206R mutation (FHR/R) share features with human atypical hemolytic uremic syndrome. Herein, we report that FHR/R mice exhibited retinal vascular occlusion and ischemia. Retinal fluorescein angiography demonstrated delayed perfusion and vascular leakage in FHR/R mice. Optical coherence tomography imaging of FHR/R mice showed retinal degeneration, edema, and detachment. Histologic analysis of FHR/R mice revealed retinal thinning, vessel occlusion, as well as degeneration of photoreceptors and retinal pigment epithelium. Immunofluorescence showed albumin leakage from blood vessels into the neural retina, and electron microscopy demonstrated vascular endothelial cell irregularity with narrowing of retinal and choroidal vessels. Knockout of C6, a component of the membrane attack complex, prevented the aforementioned retinal phenotype in FHR/R mice, consistent with membrane attack complex-mediated pathogenesis. Pharmacologic blockade of C5 also rescued retinas of FHR/R mice. This FHR/R mouse strain represents a model for retinal vascular occlusive disorders and ischemic retinopathy. The results suggest complement dysregulation can contribute to retinal vascular occlusion and that an anti-C5 antibody might be helpful for C5-mediated thrombotic retinal diseases.

PRAME as a Potential Biomarker for Liver Metastasis of Gastric Cancer.

BACKGROUND: Liver metastasis of gastric cancer (GC) is highly associated with poor prognosis. The development of sensitive biomarkers for detecting and predicting liver metastasis is required for better clinical outcome. OBJECTIVE: In this study, we aimed to identify novel genes associated with liver metastasis of GC. METHODS: Global expression profiling of 57,749 genes was performed using surgically resected gastric tissues from four patients with liver metastasis to identify candidate genes. The mRNA expression levels of the selected candidate gene were analyzed in the resected gastric tissues of 300 GC patients and correlated with clinicopathological parameters. Fourteen GC cell lines were subjected to mRNA expression and polymerase chain reaction (PCR) array analysis. RESULTS: Among 25 candidate genes identified by transcriptome analysis, preferentially expressed antigen of melanoma (PRAME) was selected for subsequent analyses. mRNA expression analysis of clinical samples revealed the aberrant expression of PRAME in GC tissues, and its high expression was significantly related to differentiated phenotype and vessel invasion, as well as liver metastasis. High PRAME expression was significantly associated with hepatic recurrence after curative surgery, and cumulative incidences of hepatic recurrence were significantly greater in patients with high PRAME expression compared with patients with low PRAME expression. In an in vitro analysis, overexpression was observed in all GC cell lines compared with a normal epithelial cell line. PCR array analysis revealed the coordinate expression of MMP9, OCLN, IL1RN, and MST1R. CONCLUSIONS: PRAME is related to the malignant potential of GC and could serve as a novel biomarker for the detection and prediction of liver metastasis.

Antibody Inhibition of Properdin Prevents Complement-Mediated Intravascular and Extravascular Hemolysis.

Paroxysmal nocturnal hemoglobinuria (PNH) is a serious blood disorder characterized by dysregulated complement activation on blood cells. Eculizumab, the current standard therapy and a humanized anti-C5 mAb, relieves anemia and thrombosis symptoms of PNH patients by preventing complement-dependent intravascular hemolysis (IVH). However, up to 20% of PNH patients on long-term eculizumab treatment still suffer from significant anemia and are transfusion dependent because of extravascular hemolysis (EVH) of C3-opsonized PNH erythrocytes. In this study, we show that function-blocking anti-properdin (P) mAbs dose-dependently inhibited autologous, complement-mediated hemolysis induced by factor H dysfunction. Furthermore, anti-human P (hP) mAbs potently and dose-dependently inhibited acidified serum-induced hemolysis of PNH erythrocytes (Ham test). In contrast to erythrocytes rescued by anti-C5 mAb, nonlysed PNH erythrocytes rescued by anti-P mAb incurred no activated C3 fragment deposition on their surface. These results suggested that anti-P mAbs may prevent EVH as well as IVH of PNH erythrocytes. To test the in vivo efficacy of anti-hP mAbs in preventing EVH, we generated a P humanized mouse by transgenic expression of hP in P knockout mice (hP-Tg/P-/-). In a murine EVH model, complement-susceptible erythrocytes were completely eliminated within 3 d in control mAb-treated hP-Tg/P-/- mice, whereas such cells were protected and persisted in hP-Tg/P-/- mice treated with an anti-hP mAb. Collectively, these data suggest that anti-P mAbs can inhibit both IVH and EVH mediated by complement and may offer improved efficacy over eculizumab, the current standard therapy for PNH.

Differential contribution of C5aR and C5b-9 pathways to renal thrombic microangiopathy and macrovascular thrombosis in mice carrying an atypical hemolytic syndrome-related factor H mutation.

Atypical hemolytic uremic syndrome (aHUS) is a form of thrombotic microangiopathy (TMA) caused by dysregulated complement activation. Clinically, aHUS is effectively treated by an anti-C5 monoclonal antibody (mAb) but whether the disease is mediated by the C5a receptor (C5aR) or C5b-9 pathway, or both, is unknown. Here we address this in a factor H mutant mouse (FHR/R) which developed complement-mediated TMA as well as macrovascular thrombosis caused by an aHUS-related factor H point mutation (mouse W1206R, corresponding to human W1183R). C5 deficiency and anti-C5 mAb treatment blocked all disease manifestations in FHR/R mice. C5aR1 gene deficiency prevented macrovascular thrombosis in various organs but did not improve survival or reduce renal TMA. Conversely, C6 or C9 deficiency significantly improved survival and markedly diminished renal TMA but did not prevent macrovascular thrombosis. Interestingly, as they aged both FHR/R C6-/- and FHR/R C9-/- mice developed glomerular disease reminiscent of C3 glomerulonephritis. Thus, C5aR and C5b-9 pathways drove different aspects of disease in FHR/R mice with the C5aR pathway being responsible for macrovascular thrombosis and chronic inflammatory injury while the C5b-9 pathway caused renal TMA. Our data provide new understanding of the pathogenesis of complement-mediated TMA and macrovascular thrombosis in FHR/R mice and suggest that C5 blockade is more effective for the treatment of aHUS than selectively targeting the C5aR or C5b-9 pathway alone.

Increased Expression of DNAJC12 is Associated with Aggressive Phenotype of Gastric Cancer.

BACKGROUND: Identification of gastric cancer-related molecules is necessary to elucidate the pathological mechanisms of this heterogeneous disease. The purpose of this study was to identify novel genes associated with aggressive phenotypes of gastric cancer. METHODS: Global expression profiling was conducted using tissues from four patients with metastatic gastric cancer to identify genes overexpressed in gastric cancer. Fifteen gastric cell lines and 262 pairs of surgically resected gastric tissues were subjected to mRNA expression analysis. The contribution of the candidate gene on gastric cancer cell proliferation, invasion, adhesion, and migration were evaluated using small interfering RNA. RESULTS: DnaJ heat shock protein family (Hsp40) member C12 (DNAJC12) was identified as a candidate gene by transcriptome analysis. In clinical samples, DNAJC12 mRNA levels were higher in gastric cancer tissues compared with normal adjacent tissues. Patients with high DNAJC12 expression showed significantly shorter overall survival in our cohort and in the extra-validation cohort analyzed by a published microarray dataset. High DNAJC12 expression in gastric cancer tissues was significantly associated with lymphatic involvement, infiltrative growth type, lymph node metastasis, and advanced stage and was identified as an independent prognostic factor for overall survival in multivariable analysis. Increased expression of DNAJC12 was found in 12 of 14 examined gastric cancer cell lines. Knockdown of DNAJC12 expression significantly decreased the proliferation and invasion abilities of gastric cancer cells. CONCLUSIONS: Our findings support DNAJC12 as a candidate gene associated with aggressive phenotypes of gastric cancer.