Rubber Ducky Cupcakes

There’s been a lot of cool science news recently, so I’ve fallen behind on posting new cupcake recipes. I thought I’d post instructions on how to make your cupcakes look like rubber ducks. (This is based on the design from the book Hello Cupcake, What’s New, which I love!) You get a huge sugar high from eating one of these, and an even bigger one if you snack on the starbursts and M&Ms that are the wrong colour, lick the frosting off your fingers, and eat the extra Timbits, marshmallows, or starburst molding failures J

Decorating Supplies:

Vanilla frosting (I use Betty Croker, do NOT use whipped)
Yellow food colouring
Mini M&M’s (brown, blue and green)
Marshmallows
Orange starbursts         
Plain Timbits
Vanilla cupcakes in yellow or white liners (start with more than you want because it takes some practice to get them looking the way you want)

 Prep Stuff:

Bake the cupcakes as described and let them cool completely.

Cut the marshmallows in half diagonally.

Cut the starbursts in half and microwave for a few seconds at a time until you can mold them into beaks (or feet). I found it helpful to make a pointy end that could be stuck into the Timbit.

Assembling the Duckies

1.      Once they have cooled, ice the cupcakes and place the marshmallow at one end and the Timbit at the other to make the head and tail of the duck. Use the frosting like glue to help hold them in place.

2.      Put them in the freezer for about 15 minutes.

3.      Microwave the frosting until it is runny and use the yellow food colouring to dye the frosting.

4.      Carefully dip the cupcake into the frosting and let excess frosting drip off.

5.      Carefully add the starburst beak and M&M eyes now OR wait until the frosting has hardened then add them at this point, whatever works out better for you. You may need to use a dab of frosting to hold the M&Ms in place. I stick the pointy end of the beak into the Timbit to help it stay in place

To make upside down duckies:

Place the marshmallow in the opposite orientation from the original ducky. Once it has been dipped, carefully place the starburst feet on either side of the marshmallow tail. You may need a small dab of icing to help them stick in place.

The placement of everything is really important, and it took several tries to get it right. The cupcakes look really good when they are done, but there were a lot of casualties in the process!

BLT Mice

As I’ve alluded to in my recent posts, HIV is kind of difficult to work with. Working with the virus in Petri dishes and cell culture flasks isn’t too bad . . . except for the fact that it’s a level 3 pathogen in Canada, which means I have to work with it in a special facility with restricted access and tons of safety features. Not all research institutions have this type of facility available, so I’m lucky to be able to do all the experiments at my research institute.

Once I complete all the studies I can do with cells, I will have to do my next group of studies using animals. This is done to test the safety in a living organism (cells can only tell me so much) and provides a more real world study system. With no idea on the safety of my compounds, I can’t do any work using humans, which is kind of a problem when studying HIV. As stated in its’ name, HIV is the HUMAN immunodeficiency virus. This means that HIV only infects humans. There are other types of immunodeficiency viruses (like simian immunodeficiency virus), but using SIV in primates, or a hybrid SIV/HIV in primates has not had good translation to work with HIV in humans, because there are a lot of differences between SIV and HIV. This has been a challenge many researchers have tried to solve, and recently success has been observed with humanized mouse models of HIV.

Before I go further I should warn you that these mice involve some very controversial stuff. First off, working with mice, which is always a touchy area. To humanize a mouse you need to perform a minor surgery on them and carefully monitor the mice post-surgery to make sure they don’t develop infections or are in pain. And, to humanize the mouse you use human fetal tissue, which can only be obtained from aborted fetuses that are donated to research. If you have any issues with any of these things, I would suggest you stop reading now.

Each humanized mouse has to be created individually. This is a very lengthy process. You begin with a mouse that, through a combination of genetic defects, does not have an immune system. There are several different types of immunodeficient mice you can use for humanization. I will be using the NOD/SCID/IL2rgamma-/- mouse. That is a fancy name for a mouse that has a severe immune deficiency and is unable to make any functional immune cells. You have to be very careful with these mice, because if they get any sort of infection their body can’t fight it so they will likely die. Their water and food is specially sterilized, their lungs are monitored for infection every other month, and sentinel mice are used to monitor for any other infections.

When the mice are 8 weeks old they have a surgery to implant human fetal liver and thymus under the kidney capsule. They also receive an IV injection of human fetal stem cells (which can be derived from the liver). The human stem cells will allow the development of all the human immune cells, and the liver/thymus allows the T cells of the immune system to mature properly. Because you use bone marrow (the stem cells), liver and thymus, these mice are called BLT mice. Twelve weeks after the surgery you take blood samples from the mice and check that all the immune cells are present. If so, the mice can be used for infection with HIV.

The BLT mice have been shown to be susceptible to vaginal infection with HIV, and the infection course is very similar to what is observed in humans. Whenever I’ve presented my research I’ve been asked how exactly you give a mouse HIV vaginally. The mouse is put under anesthesia, then a small instrument is inserted into the vagina and HIV in saline is deposited. The mouse is held in an inverted position so the saline doesn’t drip out. After a few minutes, the mouse can be woken up from anesthesia. Blood samples can be taken each week to determine HIV infection.

Today I am heading to a collaborators’ lab for 2 weeks to receive the surgical training I need to be able to make BLT mice. These mice will provide me with a lot of information about the “real world” uses of my compounds. I have to do everything perfectly for this mouse experiment, because it is really expensive. With the costs of my training trip, buying the mice, housing the mice, buying reagents and materials, and equipment rental costs, the 45-60 mouse cohort I can make from one set of donor organs will cost $20,000 or more.

Christina

PrEP for HIV Prevention

Another blog post on HIV . . . it’s sort of my thing :p (and the focus of my thesis!)

On May 10^th the Antiviral Drugs Advisory Committee of the US FDA debated whether an anti-HIV drug currently used as an antiretroviral treatment should be approved as a prevention method for uninfected people. It was recommended to the FDA that the drug, Truvada (made by Gilead Sciences), be approved for pre-exposure prophylaxis (PrEP).

PrEP means that the drug is taken routinely by uninfected people at high risk of becoming infected with HIV. It has been shown that taking the drug daily can lower risk of infection by more than 90% in two target groups – men who have sex with men; and uninfected heterosexual (men) with a long-term, HIV positive partner. However, clinical trials of the drug in heterosexual women have failed. (Also, in reading another article about this, it was claimed risk was lowered only by 42% and 73% in separate trials...)

PrEP (like birth control) would need to be taken daily to be effective. And, if adherence is low this could have disastrous consequences, particularly if a backup method of prevention (condoms) is not used. As I mentioned in my last HIV post, the virus replicates really quickly, and doesn’t make perfect copies of itself, leading to mutated forms of the virus. Some of these mutants could be resistant to Truvada. So, if you are not using PrEP as prescribed and have a partner that is HIV positive, it is likely that you will become infected with the virus. If you continue to take the PrEP irregularly, you are putting yourself at risk of being infected with drug-resistant HIV, which would likely make treatment more difficult. Of particular note is that HIV treatment with Truvada must be undertaken in combination with other antiretroviral drugs to prevent resistance.

During the clinical trials participants were monitored monthly for HIV infection, using the most sensitive test to detect HIV, so that PrEP could be stopped upon infection to prevent resistance. A few cases of infection were seen in the trial participants, and just over half of them developed resistance. This testing should be continued for those that are taking Truvada for PrEP, but this testing was not included in the recommendation to the FDA. Without regular testing, it is possible that HIV could be transmitted to someone taking PrEP (remember, it isn’t 100% effective) and they wouldn’t know about it, so they would keep taking the antiretroviral. This will lead to development of a resistant virus, which will create a lot of problems for further treatment once the virus is detected.

I did a PubMed search and wasn’t able to find any clinical trial papers about Truvada, which I would really like to be able to read before forming an opinion on this, particularly since Science and Nature report two different statistics for protection levels. I think this approval of Truvada could drastically lower HIV prevention rates, if used properly (and if affordable!), however, it could also be disastrous if used improperly (particularly without regular HIV screening). Condoms are more effective at preventing HIV transmission and I highly recommend them because they also protect against other STI’s and pregnancy. But, for those who are unable to use condoms for various reasons (particularly women in developing countries), Truvada represents a new hope for HIV prevention.

Christina

A cure for HIV?

According to the World Health Organization and UNAIDS, over 33 million people worldwide are infected with HIV, the virus that causes AIDS. Over 2 million people die AIDS-related deaths each year. HIV/AIDS has enormous social and economic impact, particularly in developing countries. Despite over 30 years of research into HIV, an effective method of prevention (besides condoms) and a cure have not been found. Currently, once someone become infected with HIV they can take a combination of anti-retroviral drugs on a strict regimen, which should suppress the virus for several years. Eventually, the patient develops AIDS which typically leads to death within 1 year. It is thought that antiretroviral therapy can increase survival time by 4-12 years, but the patient would still eventually develop AIDS.

It is really difficult to treat HIV for several reasons. The virus replicates very fast (in less than 2 days) and doesn’t always make a perfect copy when it replicates. This leads to mutations that can allow the virus to become resistant to the antiretrovirals, or to escape the immune system. All of these mutated virus also replicate quickly, so someone infected with HIV will have millions of copies of different versions of HIV. In addition, HIV infects (and kills) CD4+ T cells, which are a very important part of the immune system. So, while your immune system is trying to fight a constantly changing invader, the invader is also killing those cells that have a key role in fighting it. What is really cool is that some genetic combinations provide protection from infection or suppress HIV replication. One that is called Δ32 CCR5 deletion is present in a small percentage of Caucasians descended from Western Europeans, and also prevents infection with bubonic plague.

I have only found 1 case where someone was cured of HIV. An HIV-positive patient with acute myeloid leukemia was given a hematopoietic stem cell transplant (basically a bone marrow transplant). Since this patient had both AML and HIV there wouldn’t be very many (or maybe even any) treatment options available, so this situation provided the opportunity to try something new. The doctors decided to give them a hematopoietic stem cell transplant using cells from a donor that had the Δ32 CCR5 deletion. The transplanted cells would not be infected with HIV, and should be able to destroy any remaining HIV-infected cells in the patient. So far, HIV suppression has been observed in this patient, and they have stopped taking antiretrovirals!

Walker et al (http://jvi.asm.org/content/86/10/5719.abstract) were interested in creating a gene therapy vector that could provide protection from HIV infection. They created a gene therapy vector that would induce the Δ32 CCR5 deletion, in addition to expressing TRIM5α (shown to prevent HIV replication) and a TAR decoy (which should prevent HIV replication).

To determine if this gene therapy vector could be clinically relevant, it needs to be tested in an animal model, which is difficult for HIV. HIV is the HUMAN immunodeficiency virus, which means it can only infect humans. This has posed a huge problem for HIV researchers, but a pretty cool solution has been found – you can create mice that have a human immune system (there will be a blog about how to do this soon, because I’m going to learn how to make my own franken-mice). These mice can be infected with HIV and the infection is similar to what is observed in people.

Walker et al isolated CD34+ hematopoietic stem cells and infected them with the gene therapy vector, then injected the cells into mice. These stem cells developed into immune system cells, including the CD4+ T cells that HIV can infect. The CD4+ T cells appeared functionally normal and contained the gene therapy vector.

Once they verified everything was working as expected, they infected the mice with HIV and monitored the mice. They found that the mice that received the gene therapy vector had significantly enhanced survival of CD4+ T cells after HIV infection. However, the level of HIV virus in the blood was not changed. This suggests that the gene therapy vector was able to keep the CD4+ T cells alive, but didn’t stop HIV from replicating.

This study is a good start for using gene therapy as HIV prevention, but follow-up work on why the HIV levels in blood were unchanged should be undertaken. It is also important to realize that in this study they were sort of putting the cart before the horse – they gave the mice the gene therapy vector before HIV infection. To build on this study they should infect mice with HIV then give them a bone marrow transplant with hematopoietic stem cells that contain the gene therapy vector to see if the HIV infection is cured – it would be more clinically relevant that way. This sort of treatment would be a long way from the clinic, particularly since you’re dealing with a treatment option that could be highly controversial, since it involves stem cells and gene therapy. I am definitely going to keep my eyes on this field!

Christina

Let them eat dirt!

Many epidemiological studies have led to the hygiene hypothesis. The hygiene hypothesis proposes that exposure to microbes in early childhood can decrease susceptibility to allergies and diseases such as asthma, irritable bowel disease (IBD) and autoimmune diseases. It’s believed that this early exposure to microbes helps the immune system function properly, which prevents the “overactivation” of an immune response that occurs in the case of allergies, asthma, etc. When it “overactivates” the immune system sees something that is not dangerous (ex. dust, pollen, peanuts) as harmful, and “freaks out”, releasing all sorts of inflammatory mediators that cause symptoms from a runny nose to full-blown anaphylactic shock. In most cases, this would be annoying or inconvenient, but in some cases it can be deadly. Although a lot is known about WHAT happens when the allergen or disease is triggered, it is not known WHY. Several studies have produced observations that suggest this is all related to hygiene, specifically that exposure to microbes in early childhood lowers the chance someone will develop allergies, asthma, IBD, etc. It has been suggested that this could be related to how “clean” we are now, particularly with the use of antibacterial soaps. It has been observed that more people in developed countries have allergies/asthma/IBD/autoimmune diseases. And, within developed countries, children that are from rural areas have lower rates of allergies and asthma compared to their counterparts in cities. It wasn’t really known why this was the case, but a recent paper in Science (http://www.sciencemag.org/content/336/6080/489.full) has provided some evidence about the immune system cells that are involved.

It has been hypothesized that invariant natural killer T (iNKT) cells play an important role in the pathogenesis of ulcerative colitis and asthma. These cells recognize lipid antigens and release large amounts of proinflammatory factors (called cytokines) when they are activated. Olszak et al investigated the age-dependent regulation of iNKT cells by use of microbes in mouse models of IBD and asthma.

They worked with germ-free (GF) and specific-pathogen-free (SPF) mice. GF mice are born without the bacteria that are normally present, and are protected from any bacterial exposure over their life. SPF mice are free of certain bacteria that would cause disease, but the normal intestinal bacteria are present. They found that the numbers of iNKT cells were increased in GF mice and appeared to be stable for life. When they induced colitis in the mice they found that the GF mice were more sensitive to colitis, with more severe weight loss, pathology, and higher mortality rates compared to SPF mice. If they re-established the microbiota (all the bacteria that should be present) in the GF mice, they found that the iNKT levels and severity of colitis was not reduced. If they took pups that had a GF mother and raised them in SPF conditions, they found a complete normalization of iNKT cells and reduced susceptibility to colitis. This indicated that the microbiota present at birth has life-long effects on health. They repeated these studies with a mouse model of asthma and had very similar observations.

This study fits with the epidemiological studies that led to the hygiene hypothesis, showing that early life exposure to microbes decreases susceptibility to diseases such as IBD and asthma, where as absence of microbial exposure may have the opposite effect. This is one of the first studies that provides a mechanism to explain the epidemiological observations that compose the hygiene hypothesis.

So, I’m not saying that you should feed your kids dirt, or stop them from washing their hands before eating, but let them play in the dirt and don’t freak out if they lick their shovel – it just might lower their risk of inflammatory diseases ;p

Christina

GM Mosquitoes – the key to dengue prevention?

Dengue fever or dengue hemorrhagic fever is caused by the dengue virus. The dengue virus is only transmitted by adult mosquitoes - mostly the Aedes aegypti mosquito. There are 4 types of dengue viruses, called DENV1, DENV2, DENV3 and DENV4. If you become infected with one type of dengue virus you will likely have a mild infection and develop lifelong immunity to that virus serotype. Immunity to the other serotypes of the virus only lasts for a short time. If you get re-infected with a different serotype of the virus, that is usually when severe complications, like the hemorrhagic fever, occur. This is a huge problem with dengue because more than one serotype is usually present in a geographic location. It has been observed that if you are infected with DENV1 then get DENV2 or DENV3, or if you have DENV3 then get DENV2 you are more likely to develop dengue hemorrhagic fever.

Dengue virus is a vector borne virus, which means that there is little person-to-person spread in the absence of mosquitoes. A mosquito acquires the virus by biting an infected person. The virus can live in the mosquito but has no detrimental effects. The mosquito is infected for life and is able to transfer the virus to a person with just a single bite. There currently aren’t really any means to prevent dengue virus infection besides using pesticides, bug spray, eliminating standing water, using mosquito nets and keeping skin covered. There also really isn’t any direct treatment for the virus – usually just the symptoms, like dehydration, are treated.

In Brazil over 10 million genetically modified mosquitoes have been released over the past year to try to combat dengue virus infections. The GM mosquitoes are male and carry a gene that causes their offspring to die before reaching adulthood. Since only adult mosquitoes spread dengue virus, this should have a significant impact on spread of the virus. It has been found that currently 85% of the mosquito eggs carry the transgene, which indicates the GM males are overriding the wild population and successfully producing offspring. This should decrease the Aedes aegypti population and dengue transmission. Since relatively little time has passed since the GM mosquitoes have been released, no information is available yet on whether this is having any effects on dengue virus rates.

What is pretty cool about this is that the scientists surveyed the residents of the area where they were going to release the mosquitoes before they did anything, to find out if they were ok with the plan. They found that 90% of the residents were in favour!

If successful, this strategy could potentially eradicate dengue virus infections worldwide! The articles online (http://www.scidev.net/en/health/genomics/news/brazil-tests-gm-mosquitoes-to-fight-dengue.html) didn’t contain a lot of information and I had some questions about the uses of this GM technology. I couldn’t find any information about if these GM male mosquitoes will eventually lead to eradication of Aedes aegypti mosquitoes, or if there are concerns that other types of Aedes mosquitoes that can spread dengue would just take over transmission. I was also curious if a similar strategy might be under consideration for other mosquito-borne viruses, like West Nile virus. Finally, as with all GM organisms, there are concerns that the offspring could breed in the wild and lead to unpredictable results. It was stated that the scientists were confident no viable offspring could be produced by transgenic mosquitoes, but no explanation about this was given. All in all, very interesting, but I want more info!

C.

Storage Space Needed

No, this is not a post about how my tiny apartment has almost no storage space – it’s about something pretty amazing that’s been going on in England since 1990 (http://www.nature.com/news/children-of-the-90s-coming-of-age-1.10396)

In 1990 researchers started collecting tissues and tons of information from over 14,500 pregnant women in Bristol and surrounding area. Once the babies were born, they collected samples from the babies and a lot of information over the entire course of their life to date. They know things from how often the kids were sick to when they left home to when they hit puberty. And they have samples including placentas, blood, urine, and baby teeth. The tissue library alone has more than 1 million entries! Imagine how much storage, both physical space and databases needed to house all this information.

Combined, there is a comprehensive biological and behavioural data set for each child. This data has generated over 700 scientific papers and has led to changes in health advice for pregnant women and discovery of genetic factors for fetal growth, obesity, allergies and bone density. And there is still tons of data and samples that haven’t even been analyzed yet!

The study was started by Jean Golding, who was eager to learn how events in pregnancy and infancy affect a child’s health and development. She recruited mothers by talking on the radio and television, and sending midwives to prenatal classes and doctor’s offices. Those mothers that enrolled encompassed more than 70% of those eligible in the region during that time. The mothers filled out extensive questionnaires (more than 100 pages long!) and the children have been followed since birth. They are collecting information from as many of the fathers as possible, in hopes of being able to draw links between as many things as possible.

The first children are now 21, and some have become parents. This can lead to a new generation of subjects that can expand this study even further. However, some have called into question if the cost of collecting samples, following subjects, and storing all the data is worth it? I think it is (but I’m not a British taxpayer...) It is so difficult to get even short-term follow-up data from patients to answer research questions, that I’m just amazed by all the potential data that can be found from examining this cohort. There are so many new associations between genetics, environment or disease that could be found, that it could improve so many aspects of health care, disease screening and even behavioural indicators to identify those at risk for mental illness.

As new analysis tools are developed, more data is collected, and children of the original cohort are enrolled, this undertaking will provide more and more information that could provide so many positive changes to health care!

C

Mutant Flu – Is it trouble for you?

A conflict that I’ve been following for months has finally been concluded. In August, research groups from the US and the Netherlands submitted papers to Science and Nature respectively, detailing their experiments creating laboratory strains of H5N1 influenza A that could spread among ferrets. This was rapidly followed by a call that the papers would not be published, or published without methods or results, from the National Science Advisory Board for Biosecurity (NSABB), amid fears that these findings could be used by bioterrorists or rogue nations to start a highly lethal flu pandemic. After months of meetings between flu experts, WHO, NSABB and several other parties, it was decided that the papers would be published in full, with additional information explaining the rationale behind the experiments and the biosafety and biosecurity precautions undertaken. The first paper was published in Nature last week.

So, what’s the big deal about H5N1? What did they actually do and find in this paper? Are there any risks? Why did they do it?

Influenza viruses are named based on two proteins – hemagglutinin (the H) and neuraminidase (the N). The hemagglutinin mediates binding of the virus to target cells, and allows the virus to infect the cell. The neuraminidase is involved in releasing new viruses from infected cells, allowing the virus to spread. Different combinations of H’s and N’s make different viruses, which humans may or may not have immunity to, depending if you’ve been infected by a similar virus before (ex. If you’ve had H1N1, you likely would not get sick, or have a milder infection with H1N2). Flu viruses can “mix” together to make new H and N combinations. What makes this problematic is that some flu viruses from other species (birds or pigs) can mix with human flu viruses to make a brand new virus that can infect humans, but which we would have no prior immunity to because humans have never seen that combination before. That’s what is happening with H5N1 – it is considered an avian flu virus, but it is capable of infecting humans. Since humans have no immunity to it, there is a very high death rate (60%). H5N1 doesn’t spread well between people, which has prevented it from becoming a highly deadly pandemic. If the H5N1 virus develops a mutation that makes it able to spread between people easily, and retains the high death rate, a lot of people would die.

In the paper by Yoshihiro Kawaoka’s group they mutated the hemagglutinin of H5N1 to find viruses that should be able to infect human cells. They used sections of human trachea and lung tissues to see if the viruses could infect human cells. Those mutated viruses that were able to infect human cells were combined with gene segments from H1N1 (2009 swine flu) to generate a reassortment virus. Since H5N1 and H1N1 are circulating at the same time, and since pigs can be infected with human, avian or swine flu, it is possible that these two viruses could mix together to make a highly pathogenic virus able to spread easily between people. They used ferrets to study the transmissibility and pathogenesis of the virus, because ferrets are a widely accepted animal model for flu studies. They found that the virus needed to acquire an additional mutation to be able to replicate in ferrets. The virus needed to acquire another mutation to be able to spread efficiently between ferrets (4 mutations total required). No ferrets died from the infection. This means that although the flu virus was able to spread between ferrets, it lost its’ ability to kill those infected with it.

These experiments were carried out at the second highest biosafety level possible. I’m not familiar with this biosafety level (enhanced BSL3), but I do work at BSL3. Our facility requires a passcode and biometric scan to enter the facility. You are required to change out of all of your street clothes (including socks and underwear) and into facility scrubs, gowns, etc. All work with a virus that can transmit through the air has to be done wearing a PAPR (http://www.maxair-systems.com/700-shroud-system.html), in a biosafety cabinet. The entire facility is under negative pressure, so no air escapes at any time. Nothing leaves the facility, including people, until you are 100% sure it is no longer infectious. All infectious inventory is monitored, so it is known where every tube or cell that could have virus goes – if any cannot be accounted for, the government is contacted immediately. All of this is for the biosafety level BELOW the one the H5N1 work was done. In addition, they state that all people working with H5N1 underwent risk assessment by the United States Criminal Justice Information Services Division. It seems like this research group has done everything possible to ensure that the work is done safely, and no unauthorized people have access to the viruses. Realistically, most researchers trained in virological, immunological or microbiological techniques could probably replicate these studies, even without the methods published with the paper. But, the likelihood that those with malicious intents have access to everything necessary to replicate these studies safely is very low. Combined with the fact that the virus did not kill any of the ferrets, I believe there is little risk to the general population from these studies.

So, why do these studies? They have identified that 4 mutations can make H5N1 transmissible among ferrets. Although it is not known if these are the only 4 mutations possible to make H5N1 transmissible, surveillance programs can now monitor H5N1-infected people or poultry for these (or similar) mutations that occur in the wild. This could significantly decrease response time or lead to viral eradication that could prevent a pandemic. In addition, antivirals and vaccines against this virus can be tested and relevant stockpiles generated. This type of study is all about preparation for the worst. A small risk now could have a big effect in a future pandemic.

C.

Vegan Chocolate Cupcakes?

I decided to try my first recipe from the page-a-day calendar – the vegan chocolate cupcake. Although I don’t get the whole vegan thing (dairy cows are in pain when not milked regularly, and hens will lay eggs regardless of fertilization potential. And there seem to be a lot of options for free range, etc) I was intrigued by cupcakes made without butter and eggs. The recipe calls for applesauce and vinegar(?!) and I wasn’t sure how these cupcakes would turn out, but was curious so I whipped them up.

From the cupcake calender (and the book 500 Cupcakes, published by Sellers Publishing, Inc.)

Before I begin, I should mention that there are “fake” vegan – I wasn’t sure where to find vegan chocolate chips, so I just used normal ones. Other than that I followed the recipe exactly. I used my KitchenAid stand mixer, but you can probably mix these cupcakes by hand because the batter is very runny. The 20 minutes baking time was exactly right for my oven. The only thing that was off for this recipe was the number of cupcakes it makes. I made 1.5 dozen regular-sized cupcakes and 12 minis.

These cupcakes are pretty good. I really like the chocolate chips in them, so I might do that occasionally when making “normal” chocolate cupcakes. They are fairly moist and there is a faint apple taste which I liked. My biggest complaint with these are that they stick to the wrappers quite a bit, which is a pain, especially with the minis. Maybe cutting out some water and adding some vegetable oil would solve this?

If I had all the ingredients hanging around my kitchen (I don’t usually have applesauce, and I wouldn’t buy it special for this) I’d make these cupcakes again. They won’t replace my regular chocolate cupcakes recipe, but they gave me an idea of the cocoa/chocolate chip substitution for my regular recipe if I’m out of chocolate squares. I do wish the calendar provided calorie information because I would make the switch to the vegan recipe if it ended up being fewer calories per cupcake. I also wondered why they used a lot of sugar instead of honey, and a quick Google search showed me that there is some controversy over whether honey should be considered vegan or not.

Overall, I’d rate these cupcakes 7/10 and will add them to my cookbook.

Here’s a photo of my cupcakes. I added the icing because I had a little hanging out in my fridge, but it isn’t vegan.

UPDATE: I stored my extra cupcakes in the freezer, and after thawing the liner magically peels off!