Studies that have compared HCV-infected patients with HCV-HIV-1 infected patients have observed that there is a longer half-life of HCV, and up to ten times higher HCV RNA levels in serum or liver [2, 4]. Co-infected patients are more likely to develop cirrhosis, which may rapidly progress to acute disease . The mechanisms for these effects are unclear, but loss of immune function by HIV-1 infection is thought to be a significant contributing factor. However, most studies of HAART suggest that it does not significantly affect the levels of HCV RNA , meaning that HIV-1 may have little effect on HCV RNA production. These studies are confusing, since evidence for HCV replication is absent, and HCV RNA levels do not correlate with the levels of infectious HCV in the blood. Unfortunately there is no commonly used system to determine these levels.
Other studies that have looked at the effects of HCV on HIV-1 infected individuals have also shown conflicting results. Early studies suggest that HCV increases the rate of progression to AIDS in co-infected individuals, but others have not seen this [2, 3]. A large Swiss study showed HCV serum-positivity was associated with more likely progression to AIDS and a less likely increase of CD4+ cell count after therapy [15, 16]. Similar determinations can be made by comparing other viral infections such as HHV-6 and HIV. A strong case can be made for HHV-6 as a co-factor in the development of AIDS. Other short-term studies have not found a correlation between CD4+ cell number and negative consequences due to HCV infection [17, 18].
In vitro studies of HIV-HCV co-infected cells have been hampered by a lack of a system that can replicate both viruses. Laskuset al.  showed that HIV-1 stimulates HCV production in macrophagesin vitro. The system, however, did not support HCV replication for extended periods of time. Macrophages, although they stay alive for extended periods of time, do not synthesize DNA and replicate.
Initialin vitro studies of HIV-HHV co-infected T-cells showed that HHV-6 increased HIV-1 production and cell death . Later studies showed that HHV-6 could decrease HIV-1 production in some cell types, such as dendritic cells  and macrophages . This difference in virus production may be accounted for by differences in activities of the LTR of HIV-1.
Patients and clinically normal individuals are frequently infected with multiple viruses. It is therefore important to understand the implications of simultaneous infection by multiple viruses. Since co-infections with HIV, HCV, and HHV-6 are frequently seen in the same individual, the development of a system to study the effects of the interactions among these viruses at the level of a single cell is important. Our experience in human viruses allowed us to develop this system. We realize that the occurrence of triple infections of single cells may be a rare event, yet it also establishes the fact that immunity and dominance are also limited statistical phenomena.
Our system for growing HCV starts with the infection of macrophages obtained from human cord blood. Macrophages are distributed all over the body, and they perform specialized functions, e.g., Kupffer's cells of liver, dendritic cells of skin, astrocytes and microglial cells of the nervous system. In addition, the infection of our human neuronal precursor cells with CIMM-HCV may be similar to the infection of macrophages with HIV-1. These macrophages may become multiply infected and function as a reservoir. Other investigators have shown the presence of HCV in human brains in the post-mortem analysis [23, 24]. Similarly, HIV-1 has also been reported in the brain environment [25–27]. These observations may partly explain the diminished cognitive function, depression and fatigue in these individuals. The presence of HIV-1 and HCV in the same cell may greatly aggravate the malady.
Our TEM analysis showed HHV-6A and HCV in the same cells, with extracellular HIV-1 particles budding or adjacent to the infected cells. Since HIV-1 assembly is completed at the plasma membrane level, their presence is only seen outside the cells. The assembly and synthesis of HHV-6A begins in the nucleus, the virion acquires a membrane as it passes through the nuclear membrane, and it matures in the cytoplasm. The synthesis and maturation of HCV is presumed to occur in the cytoplasm. This, in our opinion, is an evolving concept. We have seen HCV particles in the nucleus, in the perinuclear space, in the cytoplasm, in vesicles, and outside cells. The presence of HCV particles in the perinuclear space may indicate that at least partial synthesis of this virus may occur in the nucleus. Recent suggestions that some nuclear proteins bind to HCV RNA support this possibility . The TEM results presented in Figures 6 and 7 suggest that HCV and HHV-6A may have similar or identical sites of replication. Others have proposed that HCV synthesis may occur in the perinuclear space [29, 30], which is unlikely.
Another question addressed by these experiments was to conclusively demonstrate that HCV can replicate well in T-cells. There was a difference in the level of virus replication when compared to B-cells but, as shown in the electron micrographs, it replicates well in T-cells. HCV has been shown to infect T-cells in addition to other cell types in infected individuals , so it is not surprising that significant production of HCV particles occursin vitro. The claims of levels of virus titers for many human viruses are merely claims. There are no definitive and reproducible methods for titrating biologically active HIV-1, HTLV-I, HTLV-II, HHV-6, or HCV with accuracy. We generally use the RNA levels as a relative indicator of virus production.
This study, in addition to our previous reports, supports the notion that HCV infects multiple hematopoietic cell types,viz monocytes-macrophages, B-cells, and T-cells. We have also reported that neuronal cells, endothelial cells (hepatocytes), and Kupffer's cells of liver can also be infected. Replication of HCV in liver cells is generally very low, which may be clinically relevant.
Since HCV, HIV-1, and HHV-6A can coexist in culture, none of these viruses prevent the others from infection and replication. However, these effects may be variable due to a number of factors that affect replication. This has relevance to viral protein production that may induce or produce pathology. How the viruses interact will be the subject of later work.