Newcastle disease virus-induced caspase-independent apoptosis pathway in BHK-21 cells

Background: Newcastle disease virus (NDV) is an important virus for humans. It is highly lethal in fowl and is newly identified as an oncolytic virus for cancer treatment. In vivo, NDV induces spleen, thymus, bursa, and mesenteric gland cell apoptosis, thereby causing immunosuppression. In vitro , NDV can induce apoptosis by caspase-dependent pathways including the mitochondria-mediated pathway and death receptor-mediated pathway. Methods: In this study, the major materials were baby hamster kidney (BHK-21) cells, NDV virus ( Miyadera strain; 10 3.8 TCID 50 /μl), and pan-caspase inhibiter Z-Val-Ala-DL-Asp-fluoromethylketone (z-VAD-fmk). All of the experiments used a viral infection MOI of 1. Apoptosis was confirmed using DNA fragmentation and TUNEL assay. Finally, the apoptosis-independent pathway was confirmed by western blot analysis and immunofluorescence. Results: In this study, we differentiate between the caspase group and caspase inhibition group (added 100 µM pan-caspase inhibitor [z-VAD-fmk]) in BHK-21 cells treated with NDV at 0, 12, and 24 h. In the DNA fragmentation and TUNEL assays, the apoptosis appears at 12 h and apoptosis increases over time, regardless of caspase or not. In protein level determination, the antiapoptotic protein Bcl-2 decreased over time, which is the opposite of how the proapoptotic proteins Bax, cytochrome C, Mst3, and AIF behaved. Further, using western blot and immunofluorescent staining, we checked the AIF and Endo G translocation from the mitochondria (cytoplasmic) to the nucleus. In addition to apoptosis, we found that NDV treatment of BHK-21 cells decreased actin, regardless of caspase. The actin always decreased in the NDV-treated BHK-21 cells at 12 and 24 h. Conclusions: NDV-mediated BHK-21 cell apoptosis mechanisms involve complex pathway; when in the normal state, the caspase-dependent pathway is main apoptosis pathway; when the caspase is suppressed, the BHK-21 can switch on the caspase-independent pathway by AIF, Endo G, or Mst3 to allow apoptosis to continue.

The History of Six Old Cultures of Mycobactebium Tuberculosis

Instances are recorded in the literature of mammalian -tubercle bacilli having lost their virulence completely during long cultivation on artificial media. In a paper published in 1925 I gave an account of the results of investigating two such strains which had been deposited in the National Collection of Type Cultures by Dr Nathan Raw. These strains were stated by Dr Raw to be lineal descendants of strains which had been given to him in 1906, one (human) by Prof. R. Koch, who had isolated it from the sputum of a case of human pulmonary tuberculosis, the other (bovine) by Prof. A. Calmette, who had obtained it from a mesenteric gland of a cow.

Mesenteric gland tuberculosis

The author draws attention to the important clinical significance in tbc of mesenteric adenitis, which is no less important than that of tracheo-bronchial adenitis. Causing local inflammation of peritoneum, mesenteric adenitis in tbc patients, therefore, leads to pain and intestinal disorders in the form of constipation and diarrhea.

On the Bacteriology of Asylum Dysentery in England

Since 1901, when Kruse first isolated his so-called “pseudo-dysenteric” bacilli from an institution outbreak of dysentery in Westphalia, much research on the subject has been carried out in Germany and in the United States, and the bacteriology has been well established. In this country, however, comparatively few outbreaks have been investigated, and of these only a few can be considered satisfactory. As a matter of fact, most of the early work is valueless, as non-selective media alone were used for isolating the supposed organism. Thus in 1898 Dr. Goodliffe investigated an epidemic in the Lancaster County Asylum, and came to the conclusion that it was due to a bacillus resembling B. coli, which he styled “the bacillus of ulcerative colitis.” A similar view was held by Dr. Gemmell in his monograph on the disease. Campbell, working at Rainhill in 1899, cultivated from the fæces of eight dysentery patients an organism which was almost certainly of the coli group, and regarded by him as a very likely cause of the disease. Although conversant with the writings of Chantemesse and Widal, who were the precursors of Shiga and were then actually working on B. dysenteriæ, he apparently failed to realise the significance of their investigations. In 1899 also, an outbreak at Mickleover Asylum was the subject of research by the Medical Superintendent, Dr. Legge, and Dr. Barwise, M.O.H. for Derbyshire. What was termed a “virulent” form of B. enteritidis sporogenes was here regarded as the cause of the disease, having been isolated from eight cases; but a “non-virulent” form was also found in the stools of healthy patients. All these observers, so far as I can gather, employed agar only for primary cultivation, and none of them carried out any satisfactory agglutination tests. In the first volume of Mott's Archives of Neurology, Durham published the results of some work emanating from the Claybury laboratory. He claimed that from the blood, bile and certain abdominal viscera of seven out of eight patients who had died from dysentery he had cultivated a coccus so minute that it was able to pass through the pores of a Berkefeld filter. Cultures were never made from the fæces; subcultivation in a series of generations was found impossible; and the results of agglutination and animal experiments were negative. Dr. Eyre, of Guy's Hospital, was certainly the first in Britain to show asylum researchers the way to a sounder bacteriological knowledge of the disease. Using the Conradi-Drigalski medium, in 1904 he recovered Shiga's bacillus from the fæces of four out of five Claybury patients suffering from the acute form of the disease, and post-mortem from two out of four additional cases, agglutination being obtained with the organisms, and an anti-dysenteric serum in a dilution of 1–200. As the result of further investigation, he found the bacillus of Flexner also present in several of the series, and in a private communication stated that he had detected it altogether in six out of nine acute, and fifteen out of thirty-five chronic cases, none of the latter yielding Shiga's bacillus. Hewlett (1904) tested the sera of two asylum dysentery patients with B. dysenteriæ (obtained from Flexner), and obtained a positive result in 1–50 in one case and 1–100 in the other. In a paper read before the Royal Academy of Medicine in 1905, McWeeny described a bacillus of the mannite-fermenting dysentery group which had been isolated from a severe (subsequently fatal) case in Clonmel Asylum. This organism was noteworthy, inasmuch as it produced no indol, and caused the death of several rabbits while an attempt was being made to immunise them. In the following year Candler, of Claybury, investigated the fæces of six cases suspected to be suffering from acute dysentery. “In five of them an organism resembling the Flexner type was isolated and submitted to the usual tests. The sixth was subsequently proved to have intestinal tuberculosis.” As the result of an examination of five cases occurring in one of the Sussex county asylums in 1908, Bushnell, using Conradi plates, found no dysentery bacilli present, but his report is short and incomplete. An important research was made by Aveline, Boycott and Macdonald of the Lister Institute in 1909. They obtained specimens of dysenteric fæces from Cotford Asylum, and from seventeen out of nineteen Flexner's bacillus was recovered. The spleen and mesenteric gland of a fatal case yielded it also. MacConkey's bile-salt-lactose-neutral-red agar was employed for separation of the organisms by these investigators for the first time in this work. Agglutination was obtained with patients' serum and organisms up to 1–200 and 500, and a multivalent dysentery horse-serum was shown to agglutinate in a dilution of 1–1,000, and when tested to 1–10,000, time allowed being two hours.