الفهرس 
INTRODUCTIONOnly 14 pages are availabe for public view
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Abstract
Infections with HBV and HCV are a global public health concern. chronic HBV and HCV infections are leading causes of death, with HCC and end-stage liver disease being the most common outcomes. HCC is the world’s sixth and Egypt’s fourth most frequent cancers. The availability of efficient antiviral medications that can significantly lower those risks could have an impact on the rate of hepatitis virus-related deaths. Recently, many therapeutic metabolites have entered the pharmaceutical industry to produce therapeutically effective compounds capable of fighting many deadly diseases. Fungal metabolites are a major and untapped source of novel therapeutic agents. Lectins are glycoproteins with sugar binding properties that are widely distributed in nature and extracted from various microorganisms. Because of their sugar specificity, they are used in clinical diagnosis and different therapeutic applications. Therefore, this study aimed to study antiviral activity of lectins purified from mushroom extracts against hepatitis B and C viruses.
Lectins were extracted and purified by several chromatographic steps. The purified lectins were characterized by HA and HAI assays. The concentration of lectins was determined by the Bradford assay and its molecular weight was assayed by SDS-PAGE and native gel. The effect of pH and temperature on lectins stability and activity were tested. The cytotoxicity and safety of the purified lectins were examined on PBMCs, Vero, and human hepatoma cell lines. The antiviral activity of lectins against HCV and HBV was examined. The antiviral mechanisms of lectins were studied to detect the ability of lectins to protect or neutralize virus effects or by inhibiting viral replication. These lectins were tested for their capacity to bind to viral receptors CD81 and SR-B1. They were also tested for their ability to inhibit HCV-NS3/NS4A protease and HBV-DDDP enzymes. Finally, various in-silico techniques were used to better understand the purified lectins antiviral mechanism.
The results of the present study revealed that
The purification level from the affinity column with 50 mM glycine-HCl/0.5 M NaCl buffer (pH3) elution was 41.63, 32.11 and 33.36 for ABL W, ABL B and POL, respectively.
A single band was obtained from SDS-PAGE at approximately 15, 16 and 37 kDa of the ABL W, ABL B and POL, respectively. Moreover, by using native gel, a band at 60, 60 and 80 kDa from ABL W, ABL B and POL, respectively, appeared.
ABL W and ABL B showed the highest HA of 4096 HU/ml with rabbit RBC‘s while POL showed the same hemagglutination titer with RBC‘s from human blood group O.
The HA of the purified lectins was found to be inhibited by 6.26 mM of fetuin. ABL B showed HAI with 200 mM cellobiose while POL showed inhibition of its agglutination activity with 100 mM melibiose. All the tested monosaccharides showed no effect on the HA of extracted lectins.
ABL W was stable at a wide pH range from 6-9, while ABL B was from 6-8. On the other hand, POL showed a narrow pH stability range from 6-7. ABL B showed the highest heat stability, up to 80 oC for 30 min.
Summary, Conclusion and Recommendations
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The addition of different metals, Mg2+, K2+, Ca2+, Ba2+, Zn2+, Mn2+, Al 3+ and Fe3+ to lectins showed no effect on their activity except that the hemagglutination of POL was diminished by adding a concentration higher than 6 mM of FeCl3.
These lectins showed minimal antioxidant activity compared to ascorbic acid.
Furthermore, the purified lectins showed a safe and non-cytotoxic effect on Vero and PBMC cell lines.
ABL W showed the lowest IC50 values and thus the highest activity against HCV using protection, blocking and direct virucidal mechanisms with 31.19, 3.13 and 1.48 μg/ml.
However, POL revealed high potency for inhibiting HBV replication with an IC50 value of 3.42 μg/ml. ABL W and ABL B exhibited the best activity against HBV using direct virucidal and blocking mechanisms with an IC50 of 0.60 and 0.035 μg/ml, respectively.
The purified lectins were found to inhibit the release of HBeAg and HBsAg in a concentration-dependent manner.
The binding capacity of the purified lectins to CD81 revealed that POL exhibited a significant binding percentage of 58.80%, while ABL B showed a significant blocking effect on SR-B1 with an IC50 value of 448.1 ng/ml.
Concerning the inhibitory effect of the purified lectins and SOF on HCV NS3/NS4A, SOF and POL revealed a significant high inhibitory effect with IC50 values of 822.29 and 878.72 ng/ml, respectively.
Finally, the inhibition of the HBV-DDDP was more effective with ABL B followed by POL in significant values with IC50 of 328.27 and 337.95 ng/ml, respectively.