Treatment of cartilage defects by subchondral drilling combined with covering with atelocollagen membrane induces osteogenesis in a rat model

Background

The coverage of the atelocollagen membrane at the chondral defect after subchondral drilling might improve the beneficial effects for cartilage repair because of the prevention of scattering and accumulation of cells and growth factors from bone marrow within the chondral defect. On the other hand, it might block cells and factors derived from the synovium or cause high pressure in the chondral defect, resulting in prevention of cells and growth factors gushing out from the bone marrow, which leads to disadvantages for cartilage repair.

Method

We tested this hypothesis in a 2-mm-diameter chondral defect created in the articular cartilage of the patellar groove in a rat models. Defects were left untreated, or were drilled or drilled and covered with an atelocollagen membrane; healing was evaluated by histology and gene expression analysis using real-time polymerase chain reaction and immunohistochemistry.

Results

Membrane coverage induced bone tissue ingrowth into the punched chondral defect. At 1 week, expression of TGFβ, Sox9, Runx2, osteocalcin, Col1a1, and Col2a1 in the drilling group was significantly higher than in the covering group. At 4 weeks, expressions of TGFβ, Runx2, and Col1a1 were all significantly higher in the drilling group, while Sox9, osteocalcin, and Col2a1 were significantly higher in the covering group. Immunohistochemistry demonstrated Sox9, osteocalcin, and type II collagen on the bony reparative tissue in the covering group.

Conclusions

These results suggest that the atelocollagen membrane coverage resulted in inhibition of cartilage repair.     

Involvement of Rev1 in alkylating agent‐induced loss of heterozygosity in Oryzias latipes

Translesion synthesis (TLS) polymerases mediate DNA damage bypass during replication. The TLS polymerase Rev1 has two important functions in the TLS pathway, including dCMP transferase activity and acting as a scaffolding protein for other TLS polymerases at the C‐terminus. Because of the former activity, Rev1 bypasses apurinic/apyrimidinic sites by incorporating dCMP, whereas the latter activity mediates assembly of multipolymerase complexes at the DNA lesions. We generated rev1 mutants lacking each of these two activities in Oryzias latipes (medaka) fish and analyzed cytotoxicity and mutagenicity in response to the alkylating agent diethylnitrosamine (DENA). Mutant lacking the C‐terminus was highly sensitive to DENA cytotoxicity, whereas mutant with reduced dCMP transferase activity was slightly sensitive to DENA cytotoxicity, but exhibited a higher tumorigenic rate than wild‐type fish. There was no significant difference in the frequency of DENA‐induced mutations between mutant with reduced dCMP transferase activity and wild‐type cultured cell. However, loss of heterozygosity (LOH) occurred frequently in cells with reduced dCMP transferase activity. LOH is a common genetic event in many cancer types and plays an important role on carcinogenesis. To our knowledge, this is the first report to identify the involvement of the catalytic activity of Rev1 in suppression of LOH.     

Risk factors for septic shock in acute obstructive pyelonephritis requiring emergency drainage of the upper urinary tract

Purpose

To assess the risk factors for septic shock in patients with acute obstructive pyelonephritis requiring emergency drainage of the upper urinary tract.

Methods

We retrospectively reviewed the records of 48 patients who underwent emergency drainage of the upper urinary tract for sepsis associated with acute obstructive pyelonephritis at our institute. Univariate and multivariate analyses were performed to identify the risk factors.

Results

Among 54 events of sepsis, we identified 20 events of septic shock requiring vasopressor therapy. Cases with shock were more likely than those without shock to have ureteral stone (70 vs 38 %, p = 0.024) and positive blood culture results (81 vs 28 %, p = 0.006). They received drainage significantly earlier than those without shock (1.0 vs 3.5 days, p < 0.001). Univariate analysis demonstrated that acute obstructive pyelonephritis by ureteral stone, rapid progression (the occurrence of symptoms to drainage ≤1 day), positive blood culture, leukocytopenia (<4,000/mm³), thrombocytopenia (<120,000/mm³), and prothrombin time international normalized ratio ≥1.20 were correlated with septic shock. Multivariate logistic regression analysis identified thrombocytopenia (p = 0.005) and positive blood culture (p = 0.040) as independent risk factors for septic shock.

Conclusions

Thrombocytopenia and positive blood culture were independent risk factors for septic shock in acute obstructive pyelonephritis requiring emergency drainage. Thrombocytopenia would be practically useful as a predictor of septic shock.     

Nanofibrous gelatin substrates for long-term expansion of human pluripotent stem cells

Nanofibrous gelatin substrates are suited for long-term expansion of human pluripotent stem cells (hPSCs) under feeder- and serum-free culture conditions. A combinatorial library with different sets of processing parameters was established to assess the culture performance of hPSCs on nanofibrous substrates in terms of cell adhesion and growth rate, using Matrigel as control. Then, the optimal conditions were applied to long-term expansion of hPSCs with several cell lines, showing a maintained pluripotency over more than 20 passages without introducing any abnormal chromosome. In addition, this approach allowed us to avoid enzymatic disassociation and mechanic cutting during passages, thereby promoting a better hPSC culture and long-term expansion.     

Characterization and Minimization of Block Copolypeptide Vesicle Cytotoxicity Using Different Hydrophobic Chain Lengths

The previously reported amphiphilic block copolypeptide, poly(l‐lysine)₆₀‐block‐poly(l‐leucine)₂₀ (K₆₀L₂₀), is able to form vesicles that can be manipulated to different sizes and be prepared in large quantities. This study expands upon that work by varying the length of the hydrophobic segment to optimize the vesicles so that they are monodisperse and low in toxicity. Copolypeptides with longer oligoleucine segments are found to have fewer toxic micelles, small aggregates, and unstable vesicles, and exhibit lower toxicity than vesicles formed from copolypeptides with shorter hydrophobic domains. Oligoleucine segments that are too long, however, result in rigid hydrophobic membranes that prevent the vesicular assemblies from being extruded into a monodisperse population of nanoscale vesicles.

     

Effects of Antiepileptics on Lateral Geniculate Nucleus–Kindled Seizures in Rats

The present study was undertaken to clarify the characteristics of lateral geniculate nucleus (LGN) kindling in rats, especially the efficacies of antiepileptics, in comparison with those of amygdala (AMG) kindling. Daily electrical stimulation of the LGN led to the development of a generalized convulsion (kangaroo posture and falling back) in all subjects, similar to AMG kindling. The kindling response of the LGN differed from that of the AMG in a number of respects, that is, a high after-discharge (AD) threshold, a large number of stimulations for completion of kindling, and a different pattern of electroencephalogram (EEG) development. On the other hand, the oral administration of sodium valproate, carbamazepine, clobazam, or zonisamide caused dose-dependent inhibitions of both seizure stage and AD duration of LGN-kindled seizures, whereas ethosuximide had no significant effects. In addition, seizure stage was more potently inhibited than AD duration by these antiepileptics, particularly with clobazam. In conclusion, LGN kindling possesses characteristics that are different from AMG kindling. In addition, it was demonstrated that LGN kindling is a useful model, similar to other types of limbic system kindling, for the evaluation of antiepileptics.